Synthesis of key intermediate of kras g12c inhibitor compound

ABSTRACT

The present invention relates to an improved, efficient, scalable process to prepare intermediate compounds, such as compound 5M, having the structureuseful for the synthesis of compounds that target KRAS G12C mutations, such as

CROSS-REFERENCE TO RELATED APPLICATIONS

The present application claims priority to and the benefit of U.S.Provisional Application No. 62/768,802, filed Nov. 16, 2018, which isincorporated herein by reference in its entirety for all purposes.

FIELD OF THE INVENTION

The present invention relates to an improved, efficient, scalableprocess to prepare intermediate compounds, such as compound 5M, havingthe structure

useful for the synthesis of compounds that inhibit KRAS G12C mutations.

BACKGROUND

KRAS gene mutations are common in pancreatic cancer, lungadenocarcinoma, colorectal cancer, gall bladder cancer, thyroid cancer,and bile duct cancer. KRAS mutations are also observed in about 25% ofpatients with NSCLC, and some studies have indicated that KRAS mutationsare a negative prognostic factor in patients with NSCLC. Recently,V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) mutationshave been found to confer resistance to epidermal growth factor receptor(EGFR) targeted therapies in colorectal cancer; accordingly, themutational status of KRAS can provide important information prior to theprescription of TKI therapy. Taken together, there is a need for newmedical treatments for patients with pancreatic cancer, lungadenocarcinoma, or colorectal cancer, especially those who have beendiagnosed to have such cancers characterized by a KRAS mutation, andincluding those who have progressed after chemotherapy.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 shows the crystal arrangement of composition 4a.

FIG. 2-1 shows XRPD overlay of Dione racemate Type A-E.

FIG. 2-2 shows XRPD overlay of (1S)-(−)-camphanic acid cocrystal.

FIG. 2-3 shows XRPD overlay of (+)-2, 3-dibenzoyl-D-tartaric acidcocrystal.

FIG. 2-4 shows XRPD overlay of D-(+)-malic acid cocrystal.

FIG. 2-5 shows XRPD overlay of M-Dione cocrystal at differenttemperatures.

FIG. 2-6 shows XRPD overlay of P-Dione cocrystal at differenttemperatures.

FIG. 2-7 shows XRPD overlay of M-Dione cocrystal and P-Dione cocrystalmixture at different temperatures.

FIG. 2-8 shows XRPD Overlay of Dione racemate at different temperatures(I/II).

FIG. 2-9 shows XRPD overlay of Dione racemate at different temperatures(II/II).

FIG. 2-10 shows Ternary Phase Diagram of M/P-Dione cocrystals.

FIG. 2-11 shows Ternary phase diagram of M/P-Dione.

FIG. 3-1 shows XRPD of Dione racemate Type A.

FIG. 3-2 shows TGA/DSC overlay of Dione racemate Type A.

FIG. 3-3 shows ¹H NMR spectrum of Dione racemate Type A.

FIG. 3-4 shows PLM image of Dione racemate Type A.

FIG. 3-5 shows XRPD of Dione racemate Type B.

FIG. 3-6 shows TGA/DSC overlay of Dione racemate Type B.

FIG. 3-7 shows ¹H NMR spectrum of Dione racemate Type B.

FIG. 3-8 shows XRPD of Dione racemate Type C.

FIG. 3-9 shows TGA/DSC overlay of Dione racemate Type C.

FIG. 3-10 shows ¹H NMR spectrum of Dione racemate Type C.

FIG. 3-11 shows XRPD of Dione racemate Type D.

FIG. 3-12 shows TGA/DSC overlay of Dione racemate Type D.

FIG. 3-13 shows ¹H NMR spectrum of Dione racemate Type D.

FIG. 3-14 shows XRPD of Dione racemate Type E.

FIG. 3-15 shows TGA/DSC overlay of Dione racemate Type E.

FIG. 3-16 shows ¹H NMR spectrum of Dione racemate Type E.

FIG. 3-17 shows XRPD of M-Dione cocrystal Type A.

FIG. 3-18 shows TGA/DSC overlay of M-Dione cocrystal Type A.

FIG. 3-19 shows ¹H NMR spectrum of M-Dione cocrystal Type A.

FIG. 3-20 shows XRPD of P-Dione cocrystal Type A.

FIG. 3-21 shows TGA/DSC overlay of P-Dione cocrystal Type A.

FIG. 3-22 shows ¹H NMR spectrum of P-Dione cocrystal Type A.

FIG. 3-23 shows XRPD overlay of Dione racemate forms.

FIG. 3-24 shows XRPD of competitive slurry samples.

FIG. 3-25 shows XRPD overlay of prepared P-Dione cocrystal.

FIG. 3-26 shows ¹H NMR spectrum overlay of M/P-Dione cocrystals.

FIG. 3-27 shows XRPD overlay of prepared P-Dione cocrystal.

FIG. 4-1 shows Inter-conversion diagram of M-Dione DBTA Cocrystalcrystal forms.

FIG. 5-1 shows XRPD overlay of M-dione DBTA cocrystal crystal forms(Type A E).

FIG. 5-2 shows XRPD overlay of M-dione DBTA cocrystal crystal forms(Type F K).

FIG. 5-3 shows XRPD overlay of M-dione DBTA cocrystal crystal forms(Type L Q).

FIG. 5-4 shows XRPD pattern of Type A.

FIG. 5-5 shows TGA/DSC curves of Type A.

FIG. 5-6 shows ¹H NMR of Type A.

FIG. 5-7 shows XRPD overlay of Type B.

FIG. 5-8 shows TGA/DSC curves of Type B.

FIG. 5-9 shows ¹H NMR of Type B.

FIG. 5-10 shows XRPD pattern of Type C.

FIG. 5-11 shows TGA/DSC curves of Type C.

FIG. 5-12 shows ¹H NMR of Type C.

FIG. 5-13 shows XRPD pattern of Type D.

FIG. 5-14 shows TGA/DSC curves of Type D.

FIG. 5-15 shows ¹H NMR of Type D.

FIG. 5-16 shows XRPD pattern of Type E.

FIG. 5-17 shows TGA/DSC curves of Type E.

FIG. 5-18 shows ¹H NMR of Type E.

FIG. 5-19 shows XRPD pattern of Type F.

FIG. 5-20 shows TGA/DSC curves of Type F.

FIG. 5-21 shows ¹H NMR of Type F.

FIG. 5-22 shows XRPD pattern of Type G.

FIG. 5-23 shows TGA/DSC curves of Type G.

FIG. 5-24 shows ¹H NMR of Type G.

FIG. 5-25 shows XRPD pattern of Type H.

FIG. 5-26 shows TGA/DSC curves of Type H.

FIG. 5-27 shows ¹H NMR of Type H.

FIG. 5-28 shows XRPD pattern of Type I.

FIG. 5-29 shows TGA/DSC curves of Type I.

FIG. 5-30 shows ¹H NMR of Type I.

FIG. 5-31 shows XRPD pattern of Type J.

FIG. 5-32 shows TGA/DSC curves of Type J.

FIG. 5-33 shows ¹H NMR of Type J.

FIG. 5-34 shows XRPD pattern of Type K.

FIG. 5-35 shows TGA/DSC curves of Type K.

FIG. 5-36 shows ¹H NMR of Type K.

FIG. 5-37 shows XRPD pattern of Type L.

FIG. 5-38 shows TGA/DSC curves of Type L.

FIG. 5-39 shows ¹H NMR of Type L.

FIG. 5-40 shows XRPD pattern of Type M.

FIG. 5-41 shows TGA/DSC curves of Type M.

FIG. 5-42 shows ¹H NMR of Type M.

FIG. 5-43 shows XRPD pattern of Type N.

FIG. 5-44 shows TGA/DSC curves of Type N.

FIG. 5-45 shows ¹H NMR of Type N.

FIG. 5-46 shows XRPD pattern of Type O.

FIG. 5-47 shows TGA/DSC curves of Type O.

FIG. 5-48 shows ¹H NMR of Type O.

FIG. 5-49 shows XRPD pattern of Type P.

FIG. 5-50 shows TGA/DSC curves of Type P.

FIG. 5-51 shows ¹H NMR of Type P.

FIG. 5-52 shows XRPD pattern of Type Q.

FIG. 5-53 shows TGA/DSC curves of Type Q.

FIG. 5-54 shows ¹H NMR of Type Q.

FIG. 6-1 shows HPLC of M-5 from resolution with1,3-diphenyl-3-oxopropanesulfonic acid.

FIG. 6-2 shows HPLC of 5 (P-atropisomer excess) from resolution with1,3-diphenyl-3-oxopropanesulfonic acid.

SUMMARY

The present invention relates to improved preparation of a compoundhaving the following chemical structure:

and key intermediates thereof, i.e., compositions and compoundscomprising the following chemical structures:

The present invention additional relates to a method of preparingCompound 5M having the following chemical structure

The present invention additional relates to a composition comprising astructure

DETAILED DESCRIPTION s

Definitions

Abbreviations: The following abbreviations may be used herein:

ACN Acetonitrile AcOH acetic acid aq or aq. Aqueous BOC or Boctert-butyloxy carbonyl BuOH n-butanol BuOAc Butanol acetate cpmecyclopentyl methyl ether CHCl₃ Trichloromethane DCE 1,2-dichloroethaneDABCO 1,4-diazabicyclo[2.2.2]octane DCM Dichloromethane DMAN,N-Dimethylacetamide DMAP 4-dimethylaminopyridine DME 1,2-dimethoxyethane DMF N,N-dimethylformamide DMSO dimethyl sulfoxide Dppf, DPPF ordppf 1,1′-bis(diphenylphosphino)ferrocene eq or eq. or equiv. EquivalentESI or ES electrospray ionization Et Ethyl Et₂O diethyl ether EtOAcethyl acetate EtOH ethanol g Grams h Hour H₂0 water HPLC high pressureliquid chromatography iPr Isopropyl IPA Isopropyl alcohol IPAc Isopropylacetate iPr₂NEt or DIPEA N-ethyl diisopropylamine (Hünig's base) KHMDSpotassium hexamethyldisilazide KOAc potassium acetate LDA Lithiumdiisopropylamide Lawesson's reagent2,4-bis(4-methoxyphenyl)-2,4-dithioxo-1,3,2,4- dithiadiphosphetane,2,4-Bis-(4-methoxyphenyl)- 1,3-dithia-2,4-diphosphetane 2,4-disulfide LCMS, LCMS, liquid chromatography mass spectroscopy LC-MS or LC/MS LGLeaving group (e.g., halogen, mesylate, triflate) LHMDS or LiHMDSlithium hexamethyldisilazide m/z mass divided by charge Me Methyl MeCNAcetonitrile MeOH Methanol Met Metal species for cross-coupling (e.g.,MgX, ZnX, SnR₃, SiR₃, B(OR)₂) 2-MeTHF 2-Methyltetrahydrofuran mgMilligrams min Minutes MIBK 4-Methyl-2-pentanone mL Milliliters MS massspectra MTBE Methyl tert-butyl ether n-BuLi n-butyl Lithium NaHMDSsodium hexamethyldisilazide NBS N-bromosuccinimide NCSN-chlorosuccinimide NMR nuclear magnetic resonance Pd₂(dba)₃tris(dibenzylideneacetone)dipalladium(0) Pd(dppf)Cl₂ · DCM [1,1'-Bis(diphenylphosphino)ferrocene]dichloropalladium(II), complex withdichloromethane Pd(PPh₃)₄ Tetrakis(triphenylphosphine)palladium(0) PhPhenyl PR or PG or protecting group Prot. group rbf round-bottom flaskRP-HPLC reverse phase high pressure liquid chromatography RT or rt roomtemperature sat. or satd. saturated SFC supercritical fluidchromatography SPhos Pd G3 or(2-Dicyclohexylphosphino-2′,6′-dimethoxybiphenyl) SPhos G3[2-(2′-amino-1, 1′-biphenyl)]palladium(II) methanesulfonate TBAFtetra-n-butylammonium fluoride TBTUN,N,N′,N′-Tetramethyl-O-(benzotriazol-1-yl)uronium tetrafluoroboratet-BuOH tert-butanol TEA or Et₃N Trimethylamine TFA trifluoroacetic acidTHF Tetrahydrofuran UV Ultraviolet XRPD X-Ray Powder Diffraction

The use of the terms “a,” “an,” “the,” and similar referents in thecontext of describing the invention (especially in the context of theclaims) are to be construed to cover both the singular and the plural,unless otherwise indicated. Recitation of ranges of values herein merelyare intended to serve as a shorthand method of referring individually toeach separate value falling within the range, unless otherwise indicatedherein, and each separate value is incorporated into the specificationas if it were individually recited herein. The use of any and allexamples, or exemplary language (e.g., “such as”) provided herein, isintended to better illustrate the invention and is not a limitation onthe scope of the invention unless otherwise claimed. No language in thespecification should be construed as indicating any non-claimed elementas essential to the practice of the invention.

As used herein, the term “alkyl” refers to straight chained and branchedC1-C₈ hydrocarbon groups, including but not limited to, methyl, ethyl,n-propyl, i-propyl, n-butyl, sec-butyl, t-butyl, n-pentyl,2-methylbutyl, 3-methylbutyl, 2,2-dimethylpropyl, n-hexyl,2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,2-dimethylbutyl,2,3-dimethylbutyl, 3,3-dimethylbutyl, and 2-ethybutyl. The term C_(m-n)means the alkyl group has “m” to “n” carbon atoms. The term “alkylene”refers to an alkyl group having a substituent. An alkyl (e.g., methyl),or alkylene (e.g., —CH₂—), group can be substituted with one or more,and typically one to three, of independently selected, for example,halo, trifluoromethyl, trifluoromethoxy, hydroxy, alkoxy, nitro, cyano,alkylamino, C₁₋₈alkyl, C₂₋₈alkenyl, C₂₋₈alkynyl, —NC, amino, —CO₂H,—CO₂C₁—C₈alkyl, —OCOC₁—C₈alkyl, C₃-C₁₀ cycloalkyl, C₃-C₁₀heterocycloalkyl, C₅-C₅aryl, and C₅-C₁₀ heteroaryl. The term “haloalkyl”specifically refers to an alkyl group wherein at least one, e.g., one tosix, or all of the hydrogens of the alkyl group are substituted withhalo atoms.

The terms “alkenyl” and “alkynyl” indicate an alkyl group that furtherincludes a double bond or a triple bond, respectively.

As used herein, the term “halo” refers to fluoro, chloro, bromo, andiodo. The term “alkoxy” is defined as —OR, wherein R is alkyl.

As used herein, the term “amino” or “amine” interchangeably refers to a—NR₂ group, wherein each R is, e.g., H or a substituent. In someembodiments, the amino group is further substituted to form an ammoniumion, e.g., NR₃ ⁺. Ammonium moieties are specifically included in thedefinition of “amino” or “amine.” Substituents can be, for example, analkyl, alkoxy, cycloalkyl, heterocycloalkyl, amide, or carboxylate. An Rgroup may be further substituted, for example, with one or more, e.g.,one to four, groups selected from halo, cyano, alkenyl, alkynyl, alkyl,cycloalkyl, heterocycloalkyl, aryl, heteroaryl, urea, carbonyl,carboxylate, amine, and amide. An “amide” or “amido” groupinterchangeably refers to a group similar to an amine or amino group butfurther including a C(O), e.g., —C(O)NR₂.

As used herein, the term “aryl” refers to a C₆₋₁₄ monocyclic orpolycyclic aromatic group, preferably a C₆₋₁₀ monocyclic or bicyclicaromatic group, or C₁₀₋₁₄ polycyclic aromatic group. Examples of arylgroups include, but are not limited to, phenyl, naphthyl, fluorenyl,azulenyl, anthryl, phenanthryl, pyrenyl, biphenyl, and terphenyl. Arylalso refers to C₁₀₋₁₄ bicyclic and tricyclic carbon rings, where onering is aromatic and the others are saturated, partially unsaturated, oraromatic, for example, dihydronaphthyl, indenyl, indanyl, ortetrahydronaphthyl (tetralinyl). Unless otherwise indicated, an arylgroup can be unsubstituted or substituted with one or more, and inparticular one to four, groups independently selected from, for example,halo, C₁₋₈alkyl, C₂₋₈alkenyl, C₂₋₈alkynyl, —CF₃, —OCF₃, —NO₂, —CN, —NC,—OH, alkoxy, amino, —CO₂H, —CO₂C₁-C₈alkyl, —OCOC₁-C₈alkyl, C₃-C₁₀cycloalkyl, C₃-C₁₀ heterocycloalkyl, C₅-C₁₀ aryl, and C₅-C₁₀ heteroaryl.

As used herein, the term “cycloalkyl” refers to a monocyclic orpolycyclic non-aromatic carbocyclic ring, where the polycyclic ring canbe fused, bridged, or spiro. The carbocyclic ring can have 3 to 10carbon ring atoms. Contemplated carbocyclic rings include, but are notlimited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl,cycloheptyl, cyclooctyl, and cyclononyl.

As used herein, the term “heterocycloalkyl” means a monocyclic orpolycyclic (e.g., bicyclic), saturated or partially unsaturated, ringsystem containing 3 or more (e.g., 3 to 12, 4 to 10, 4 to 8, or 5 to 7)total atoms, of which one to five (e.g., 1, 2, 3, 4, or 5) of the atomsare independently selected from nitrogen, oxygen, and sulfur.Nonlimiting examples of heterocycloalkyl groups include azetidinyl,pyrrolidinyl, piperidinyl, piperazinyl, dihydropyrrolyl, morpholinyl,thiomorpholinyl, dihydropyridinyl, oxacycloheptyl, dioxacycloheptyl,thiacycloheptyl, and diazacycloheptyl.

Unless otherwise indicated, a cycloalkyl or heterocycloalkyl group canbe unsubstituted or substituted with one or more, and in particular oneto four, groups. Some contemplated substituents include halo, C₁₋₈alkyl,C₂₋₈alkenyl, C₂₋₈alkynyl, —OCF₃, —NO₂, —CN, —NC, —OH, alkoxy, amino,—CO₂H, —CO₂C₁-C₈alkyl, —OCOC₁-C₈alkyl, C₃-C₁₀ cycloalkyl, C₃-C₁₀heterocycloalkyl, C₅-C₁₀aryl, and C₅-C₁₀ heteroaryl.

As used herein, the term “heteroaryl” refers to a monocyclic orpolycyclic ring system (for example, bicyclic) containing one to threearomatic rings and containing one to four (e.g., 1, 2, 3, or 4)heteroatoms selected from nitrogen, oxygen, and sulfur in an aromaticring. In certain embodiments, the heteroaryl group has from 5 to 20,from 5 to 15, from 5 to 10 ring, or from 5 to 7 atoms. Heteroaryl alsorefers to C10-14 bicyclic and tricyclic rings, where one ring isaromatic and the others are saturated, partially unsaturated, oraromatic. Examples of heteroaryl groups include, but are not limited to,furanyl, imidazolyl, isothiazolyl, isoxazolyl, oxadiazolyl, oxazolyl,pyrazinyl, pyrazolyl, pyridazinyl, pyridyl, pyrimidinyl, pyrrolyl,thiadiazolyl, thiazolyl, thienyl, tetrazolyl, triazinyl, triazolyl,benzofuranyl, benzimidazolyl, benzoisoxazolyl, benzopyranyl,benzothiadiazolyl, benzothiazolyl, benzothienyl, benzothiophenyl,benzotriazolyl, benzoxazolyl, furopyridyl, imidazopyridinyl,imidazothiazolyl, indolizinyl, indolyl, indazolyl, isobenzofuranyl,isobenzothienyl, isoindolyl, isoquinolinyl, isothiazolyl,naphthyridinyl, oxazolopyridinyl, phthalazinyl, pteridinyl, purinyl,pyridopyridyl, pyrrolopyridyl, quinolinyl, quinoxalinyl, quiazolinyl,thiadiazolopyrimidyl, and thienopyridyl. Unless otherwise indicated, aheteroaryl group can be unsubstituted or substituted with one or more,and in particular one to four or one or two, substituents. Contemplatedsubstituents include halo, C₁₋₈alkyl, C₂₋₈alkenyl, C₂₋₈alkynyl, —OCF₃,—NO₂, —CN, —NC, —OH, alkoxy, amino, —CO₂H, —CO₂C₁-C₈alkyl,—OCOC₁-C₈alkyl, C₃-C₁₀ cycloalkyl, C₃-C₁₀ heterocycloalkyl, C5-C₁₀ aryl,and C5-C₁₀ heteroaryl.

As used herein, the term Boc refers to the structure

EMBODIMENTS Embodiment 1

In one embodiment of the invention, the present invention comprises acomposition, the composition comprising a compound of Formula 4:

and a compound of Formula B:

Embodiment 2

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 1, wherein the compound ofFormula 4 is a compound of Formula 5M:

Embodiment 3

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 1, wherein the compound ofFormula 4 is a compound of Formula 5P:

Emb 4

In another embodiment of the present invention, the present inventioncomprises the composition of any one of embodiments 1-3, wherein thecompound of Formula B is a compound of formula B1:

Embodiment 5

In another embodiment of the present invention, the present inventioncomprises the composition of any one of embodiments 1-3, wherein thecompound of Formula B is a compound of formula B2:

Embodiment 6

In another embodiment of the present invention, the present inventioncomprises the composition of any one of embodiments 1-5, wherein thecomposition comprises a 2 to 1 ratio of the compound of Formula 4 to thecompound of Formula B.

Embodiment 7

In another embodiment of the present invention, the present inventioncomprises the composition of any one of embodiments 1-6, wherein thecomposition further comprises 2-methyltetrahydrofuran having theformula:

Embodiment 8

In another embodiment of the present invention, the present inventioncomprises the composition of any one of embodiments 1-7, wherein theratio of the 2-methyltetrahydrofuran to the compound of formula B is 2to 1.

Embodiment 9

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 1, wherein the composition hasthe formula:

Embodiment 10

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 9, wherein the composition hasthe formula:

Embodiment 11

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 9, wherein the composition hasthe formula:

Embodiment 12

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 9, wherein the composition hasthe formula:

Embodiment 13

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 9, wherein the composition hasthe formula:

Embodiment 14

In another embodiment of the present invention, the present inventioncomprises the composition of any one of embodiments 1-13, wherein thecomposition is in a crystalline state.

Embodiment 15

In another embodiment of the present invention, the present inventioncomprises a method of making a composition of formula 4a, the methodcomprising reacting a compound 4, having the following chemicalstructure:

with a compound B1, having the formula:

in the presence of 2-methyltetrahydrofuran to form the composition offormula 4a, having the structure:

Embodiment 16

In another embodiment of the present invention, the present inventioncomprises a method of obtaining a compound of formula 5M, having thefollowing chemical structure:

the method comprising:

-   a) reacting a compound 4, having the following chemical structure:

with a compound B1, having the formula:

in the presence of 2-methyltetrahydrofuran to form a composition offormula 4a, having the structure:

as crystals;

-   b) isolating composition 4a, and-   c) treating the isolated composition 4a with a base to produce the    compound of formula 5M.

Embodiment 17

In another embodiment of the present invention, the present inventioncomprises the method according to Embodiment 16, wherein the base isNa₂HPO₄.

Embodiment 18

In another embodiment of the present invention, the present inventioncomprises the method according to Embodiment 16, wherein the base isNaHCO₃.

Embodiment 19

In another embodiment of the present invention, the present inventioncomprises the composition, the composition comprising a compound ofFormula 4:

and a compound of Formula 11:

Embodiment 20

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 19, wherein the compound ofFormula 4 is a compound of Formula 5M:

Embodiment 21

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 19, wherein the compound ofFormula 4 is a compound of Formula 5P:

Embodiment 22

In another embodiment of the present invention, the present inventioncomprises the composition of any one of embodiments 19-21, wherein thecompound of Formula 11 is a compound of formula 11 a:

Embodiment 23

In another embodiment of the present invention, the present inventioncomprises the composition of any one of embodiments 19-21, wherein thecompound of Formula 11 is a compound of formula 11b:

Embodiment 24

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 19, wherein the composition hasthe formula:

Embodiment 25

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 19, wherein the composition hasthe formula:

Embodiment 26

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 19, wherein the composition hasthe formula:

Embodiment 27

In another embodiment of the present invention, the present inventioncomprises the composition of embodiment 19, wherein the composition hasthe formula:

Embodiment 28

In another embodiment of the present invention, the present inventioncomprises the composition of any one of embodiments 19-27, wherein thecomposition comprises a 1 to 1 ratio of the compound of Formula 4 to thecompound of Formula 11.

Embodiment 29

In another embodiment of the present invention, the present inventioncomprises the method of embodiment 16, wherein the compound of formula5M is used as an intermediate to generate a compound having the Formula9:

Embodiment 30

The method of embodiment 29, wherein the method further comprises mixingthe compound of Formula 9 with at least one pharmaceutically acceptableexcipient to form a pharmaceutical composition.

Compounds of the disclosure

Provided herein are KRAS inhibitors having structures discussed in moredetail below.

The compounds disclosed herein include all pharmaceutically acceptableisotopically-labeled compounds wherein one or more atoms of thecompounds disclosed herein are replaced by atoms having the same atomicnumber, but an atomic mass or mass number different from the atomic massor mass number usually found in nature. Examples of isotopes that can beincorporated into the disclosed compounds include isotopes of hydrogen,carbon, nitrogen, oxygen, phosphorous, fluorine, chlorine, and iodine,such as ²H, ³H, ¹¹C, ¹³C, ¹⁴C, ¹³N, ¹⁵N, ¹⁵O, ¹⁷O, ¹⁸O, ³¹F, ³²F, ³⁵S,¹⁸F, ³⁶Cl, ¹²³I, and ¹²⁵I, respectively. These radiolabelled compoundscould be useful to help determine or measure the effectiveness of thecompounds, by characterizing, for example, the site or mode of action,or binding affinity to pharmacologically important site of action.Certain isotopically-labeled compounds of the disclosure, for example,those incorporating a radioactive isotope, are useful in drug and/orsubstrate tissue distribution studies. The radioactive isotopes tritium,i.e. ³H, and carbon-14, i.e. ¹⁴C, are particularly useful for thispurpose in view of their ease of incorporation and ready means ofdetection.

Substitution with heavier isotopes such as deuterium, i.e. ²H, mayafford certain therapeutic advantages resulting from greater metabolicstability, for example, increased in vivo half-life or reduced dosagerequirements, and hence are preferred in some circumstances.

Substitution with positron emitting isotopes, such as ¹¹C, ¹⁸F, ¹⁵O and¹³N, can be useful in Positron Emission Topography (PET) studies forexamining substrate receptor occupancy. Isotopically-labeled compoundsof structure (I) can generally be prepared by conventional techniquesknown to those skilled in the art or by processes analogous to thosedescribed in the Preparations and Examples as set out below using anappropriate isotopically-labeled reagent in place of the non-labeledreagent previously employed.

Isotopically-labeled compounds as disclosed herein can generally beprepared by conventional techniques known to those skilled in the art orby processes analogous to those described in the accompanying examplesand schemes using an appropriate isotopically-labeled reagent in placeof the non-labeled reagent previously employed.

Certain of the compounds as disclosed herein may exist as stereoisomers(i.e., isomers that differ only in the spatial arrangement of atoms)including optical isomers and conformational isomers (or conformers).The compounds disclosed herein include all stereoisomers, both as pureindividual stereoisomer preparations and enriched preparations of each,and both the racemic mixtures of such stereoisomers as well as theindividual diastereomers and enantiomers that may be separated accordingto methods that are known to those skilled in the art. Additionally, thecompounds disclosed herein include all tautomeric forms of thecompounds.

Certain of the compounds disclosed herein may exist as atropisomers,which are conformational stereoisomers that occur when rotation about asingle bond in the molecule is prevented, or greatly slowed, as a resultof steric interactions with other parts of the molecule. The compoundsdisclosed herein include all atropisomers, both as pure individualatropisomer preparations, enriched preparations of each, or anon-specific mixture of each. Where the rotational barrier about thesingle bond is high enough, and interconversion between conformations isslow enough, separation and isolation of the isomeric species may bepermitted. For example, groups such as, but not limited to, thefollowing groups

may exhibit restricted rotation.

The term “monohydrate” means a salt of Compound 9 having about oneassociated water molecule. Those skilled in the art appreciate that theexact number of the associated water molecules may vary slightly at anytime with variable temperature, pressure, and other environmentalinfluence. All slight variations of the number of the associated watermolecules are contemplated to be within the scope of the presentinvention.

The term “dihydrate” means a salt of Compound 9 having about twoassociated water molecules. Those skilled in the art appreciate that theexact number of the associated water molecules may vary slightly at anytime with variable temperature, pressure, and other environmentalinfluence. All slight variations of the number of the associated watermolecules are contemplated to be within the scope of the presentinvention.

The term “co-crystal” means a crystalline material comprising two ormore compounds at ambient temperature (20° C. to 25° C., preferably 20°C.), of which at least two are held together by weak interaction,wherein at least one of the compounds is a co-crystal former and theother is Compound 5. Weak interaction is being defined as an interactionwhich is neither ionic nor covalent and includes for example: hydrogenbonds, van der Waals forces, and 7E-7E interactions.

The term “amorphous form” or “amorphous” means a material that lackslong range order and as such does not show distinct X-ray diffractionpeaks, i.e. a Bragg diffraction peak. The XRPD pattern of an amorphousmaterial is characterized by one or more amorphous halos.

The term “amorphous halo” is an approximately bell-shaped maximum in theX-ray powder pattern of an amorphous substance.

The term “substantially pure” refers to a solid form of Compound 9having purity greater than about 95%, specifically greater than about99.5%, more specifically greater than about 99.8% and still morespecifically greater than about 99.9%.

The term “patient” means animals, such as dogs, cats, cows, horses,sheep and humans. Particular patients are mammals. The term patientincludes males and females.

The terms “treating”, “treat” or “treatment” and the like includepreventative (e.g., prophylactic) and palliative treatment.

The term “excipient” means any pharmaceutically acceptable additive,carrier, diluent, adjuvant, or other ingredient, other than the activepharmaceutical ingredient (API), which is typically included forformulation and/or administration to a patient.

Pharmaceutical Compositions, Dosing, and Routes of Administration

Also provided herein are pharmaceutical compositions that include acompound as disclosed herein, together with a pharmaceuticallyacceptable excipient, such as, for example, a diluent or carrier.Compounds and pharmaceutical compositions suitable for use in thepresent invention include those wherein the compound can be administeredin an effective amount to achieve its intended purpose. Administrationof the compound described in more detail below.

Suitable pharmaceutical formulations can be determined by the skilledartisan depending on the route of administration and the desired dosage.See, e.g., Remington's Pharmaceutical Sciences, 1435-712 (18th ed., MackPublishing Co, Easton, Pa., 1990). Formulations may influence thephysical state, stability, rate of in vivo release and rate of in vivoclearance of the administered agents. Depending on the route ofadministration, a suitable dose may be calculated according to bodyweight, body surface areas or organ size. Further refinement of thecalculations necessary to determine the appropriate treatment dose isroutinely made by those of ordinary skill in the art without undueexperimentation, especially in light of the dosage information andassays disclosed herein as well as the pharmacokinetic data obtainablethrough animal or human clinical trials.

The phrases “pharmaceutically acceptable” or “pharmacologicallyacceptable” refer to molecular entities and compositions that do notproduce adverse, allergic, or other untoward reactions when administeredto an animal or a human. As used herein, “pharmaceutically acceptable”includes any and all solvents, dispersion media, coatings, antibacterialand antifungal agents, isotonic and absorption delaying agents and thelike. The use of such excipients for pharmaceutically active substancesis well known in the art. Except insofar as any conventional media oragent is incompatible with the therapeutic compositions, its use intherapeutic compositions is contemplated. Supplementary activeingredients also can be incorporated into the compositions. In exemplaryembodiments, the formulation may comprise corn syrup solids, high-oleicsafflower oil, coconut oil, soy oil, L-leucine, calcium phosphatetribasic, L-tyrosine, L-proline, L-lysine acetate, DATEM (anemulsifier), L-glutamine, L-valine, potassium phosphate dibasic,L-isoleucine, L-arginine, L-alanine, glycine, L-asparagine monohydrate,L-serine, potassium citrate, L-threonine, sodium citrate, magnesiumchloride, L-histidine, L-methionine, ascorbic acid, calcium carbonate,L-glutamic acid, L-cystine dihydrochloride, L-tryptophan, L-asparticacid, choline chloride, taurine, m-inositol, ferrous sulfate, ascorbylpalmitate, zinc sulfate, L-carnitine, alpha-tocopheryl acetate, sodiumchloride, niacinamide, mixed tocopherols, calcium pantothenate, cupricsulfate, thiamine chloride hydrochloride, vitamin A palmitate, manganesesulfate, riboflavin, pyridoxine hydrochloride, folic acid,beta-carotene, potassium iodide, phylloquinone, biotin, sodium selenate,chromium chloride, sodium molybdate, vitamin D3 and cyanocobalamin.

The compound can be present in a pharmaceutical composition as apharmaceutically acceptable salt. As used herein, “pharmaceuticallyacceptable salts” include, for example base addition salts and acidaddition salts.

Pharmaceutically acceptable base addition salts may be formed withmetals or amines, such as alkali and alkaline earth metals or organicamines. Pharmaceutically acceptable salts of compounds may also beprepared with a pharmaceutically acceptable cation. Suitablepharmaceutically acceptable cations are well known to those skilled inthe art and include alkaline, alkaline earth, ammonium and quaternaryammonium cations. Carbonates or hydrogen carbonates are also possible.Examples of metals used as cations are sodium, potassium, magnesium,ammonium, calcium, or ferric, and the like. Examples of suitable aminesinclude isopropylamine, trimethylamine, histidine,N,N′-dibenzylethylenediamine, chloroprocaine, choline, diethanolamine,dicyclohexylamine, ethylenediamine, N-methylglucamine, and procaine.

Pharmaceutically acceptable acid addition salts include inorganic ororganic acid salts. Examples of suitable acid salts include thehydrochlorides, formates, acetates, citrates, salicylates, nitrates,phosphates. Other suitable pharmaceutically acceptable salts are wellknown to those skilled in the art and include, for example, formic,acetic, citric, oxalic, tartaric, or mandelic acids, hydrochloric acid,hydrobromic acid, sulfuric acid or phosphoric acid; with organiccarboxylic, sulfonic, sulfo or phospho acids or N-substituted sulfamicacids, for example acetic acid, trifluoroacetic acid (TFA), propionicacid, glycolic acid, succinic acid, maleic acid, hydroxymaleic acid,methylmaleic acid, fumaric acid, malic acid, tartaric acid, lactic acid,oxalic acid, gluconic acid, glucaric acid, glucuronic acid, citric acid,benzoic acid, cinnamic acid, mandelic acid, salicylic acid,4-aminosalicylic acid, 2-phenoxybenzoic acid, 2-acetoxybenzoic acid,embonic acid, nicotinic acid or isonicotinic acid; and with amino acids,such as the 20 alpha amino acids involved in the synthesis of proteinsin nature, for example glutamic acid or aspartic acid, and also withphenylacetic acid, methanesulfonic acid, ethanesulfonic acid,2-hydroxyethanesulfonic acid, ethane 1,2-disulfonic acid,benzenesulfonic acid, 4-methylbenzenesulfonic acid, naphthalene2-sulfonic acid, naphthalene 1,5-disulfonic acid, 2- or3-phosphoglycerate, glucose 6-phosphate, N-cyclohexylsulfamic acid (withthe formation of cyclamates), or with other acid organic compounds, suchas ascorbic acid.

Pharmaceutical compositions containing the compounds disclosed hereincan be manufactured in a conventional manner, e.g., by conventionalmixing, dissolving, granulating, dragee-making, levigating, emulsifying,encapsulating, entrapping, or lyophilizing processes. Proper formulationis dependent upon the route of administration chosen.

For oral administration, suitable compositions can be formulated readilyby combining a compound disclosed herein with pharmaceuticallyacceptable excipients such as carriers well known in the art. Suchexcipients and carriers enable the present compounds to be formulated astablets, pills, dragees, capsules, liquids, gels, syrups, slurries,suspensions and the like, for oral ingestion by a patient to be treated.Pharmaceutical preparations for oral use can be obtained by adding acompound as disclosed herein with a solid excipient, optionally grindinga resulting mixture, and processing the mixture of granules, afteradding suitable auxiliaries, if desired, to obtain tablets or drageecores. Suitable excipients include, for example, fillers and cellulosepreparations. If desired, disintegrating agents can be added.Pharmaceutically acceptable ingredients are well known for the varioustypes of formulation and may be for example binders (e.g., natural orsynthetic polymers), lubricants, surfactants, sweetening and flavoringagents, coating materials, preservatives, dyes, thickeners, adjuvants,antimicrobial agents, antioxidants and carriers for the variousformulation types.

When a therapeutically effective amount of a compound disclosed hereinis administered orally, the composition typically is in the form of asolid (e.g., tablet, capsule, pill, powder, or troche) or a liquidformulation (e.g., aqueous suspension, solution, elixir, or syrup).

When administered in tablet form, the composition can additionallycontain a functional solid and/or solid carrier, such as a gelatin or anadjuvant. The tablet, capsule, and powder can contain about 1 to about95% compound, and preferably from about 15 to about 90% compound.

When administered in liquid or suspension form, a functional liquidand/or a liquid carrier such as water, petroleum, or oils of animal orplant origin can be added. The liquid form of the composition canfurther contain physiological saline solution, sugar alcohol solutions,dextrose or other saccharide solutions, or glycols. When administered inliquid or suspension form, the composition can contain about 0.5 toabout 90% by weight of a compound disclosed herein, and preferably about1 to about 50% of a compound disclosed herein. In one embodimentcontemplated, the liquid carrier is non-aqueous or substantiallynon-aqueous. For administration in liquid form, the composition may besupplied as a rapidly-dissolving solid formulation for dissolution orsuspension immediately prior to administration.

When a therapeutically effective amount of a compound disclosed hereinis administered by intravenous, cutaneous, or subcutaneous injection,the composition is in the form of a pyrogen-free, parenterallyacceptable aqueous solution. The preparation of such parenterallyacceptable solutions, having due regard to pH, isotonicity, stability,and the like, is within the skill in the art. A preferred compositionfor intravenous, cutaneous, or subcutaneous injection typicallycontains, in addition to a compound disclosed herein, an isotonicvehicle. Such compositions may be prepared for administration assolutions of free base or pharmacologically acceptable salts in watersuitably mixed with a surfactant, such as hydroxypropylcellulose.Dispersions also can be prepared in glycerol, liquid polyethyleneglycols, and mixtures thereof and in oils. Under ordinary conditions ofstorage and use, these preparations can optionally contain apreservative to prevent the growth of microorganisms.

Injectable compositions can include sterile aqueous solutions,suspensions, or dispersions and sterile powders for the extemporaneouspreparation of sterile injectable solutions, suspensions, ordispersions. In all embodiments the form must be sterile and must befluid to the extent that easy syringability exists. It must be stableunder the conditions of manufacture and storage and must resist thecontaminating action of microorganisms, such as bacteria and fungi, byoptional inclusion of a preservative. The carrier can be a solvent ordispersion medium containing, for example, water, ethanol, polyol (e.g.,glycerol, propylene glycol, and liquid polyethylene glycol, and thelike), suitable mixtures thereof, and vegetable oils. In one embodimentcontemplated, the carrier is non-aqueous or substantially non-aqueous.The proper fluidity can be maintained, for example, by the use of acoating, such as lecithin, by the maintenance of the required particlesize of the compound in the embodiment of dispersion and by the use ofsurfactants. The prevention of the action of microorganisms can bebrought about by various antibacterial and antifungal agents, forexample, parabens, chlorobutanol, phenol, sorbic acid, thimerosal, andthe like. In many embodiments, it will be preferable to include isotonicagents, for example, sugars or sodium chloride. Prolonged absorption ofthe injectable compositions can be brought about by the use in thecompositions of agents delaying absorption, for example, aluminummonostearate and gelatin.

Sterile injectable solutions are prepared by incorporating the activecompounds in the required amount in the appropriate solvent with variousof the other ingredients enumerated above, as required, followed byfiltered sterilization. Generally, dispersions are prepared byincorporating the various sterilized active ingredients into a sterilevehicle which contains the basic dispersion medium and the requiredother ingredients from those enumerated above. In the embodiment ofsterile powders for the preparation of sterile injectable solutions, thepreferred methods of preparation are vacuum-drying and freeze-dryingtechniques which yield a powder of the active ingredient plus anyadditional desired ingredient from a previously sterile-filteredsolution thereof.

Slow release or sustained release formulations may also be prepared inorder to achieve a controlled release of the active compound in contactwith the body fluids in the GI tract, and to provide a substantiallyconstant and effective level of the active compound in the blood plasma.For example, release can be controlled by one or more of dissolution,diffusion, and ion-exchange. In addition, the slow release approach mayenhance absorption via saturable or limiting pathways within the GItract. For example, the compound may be embedded for this purpose in apolymer matrix of a biological degradable polymer, a water-solublepolymer or a mixture of both, and optionally suitable surfactants.Embedding can mean in this context the incorporation of micro-particlesin a matrix of polymers. Controlled release formulations are alsoobtained through encapsulation of dispersed micro-particles oremulsified micro-droplets via known dispersion or emulsion coatingtechnologies.

For administration by inhalation, compounds of the present invention areconveniently delivered in the form of an aerosol spray presentation frompressurized packs or a nebulizer, with the use of a suitable propellant.In the embodiment of a pressurized aerosol, the dosage unit can bedetermined by providing a valve to deliver a metered amount. Capsulesand cartridges of, e.g., gelatin, for use in an inhaler or insufflatorcan be formulated containing a powder mix of the compound and a suitablepowder base such as lactose or starch.

The compounds disclosed herein can be formulated for parenteraladministration by injection (e.g., by bolus injection or continuousinfusion). Formulations for injection can be presented in unit dosageform (e.g., in ampules or in multidose containers), with an addedpreservative. The compositions can take such forms as suspensions,solutions, or emulsions in oily or aqueous vehicles, and can containformulatory agents such as suspending, stabilizing, and/or dispersingagents.

Pharmaceutical formulations for parenteral administration includeaqueous solutions of the compounds in water-soluble form. Additionally,suspensions of the compounds can be prepared as appropriate oilyinjection suspensions. Suitable lipophilic solvents or vehicles includefatty oils or synthetic fatty acid esters. Aqueous injection suspensionscan contain substances which increase the viscosity of the suspension.Optionally, the suspension also can contain suitable stabilizers oragents that increase the solubility of the compounds and allow for thepreparation of highly concentrated solutions. Alternatively, a presentcomposition can be in powder form for constitution with a suitablevehicle (e.g., sterile pyrogen-free water) before use.

Compounds disclosed herein also can be formulated in rectalcompositions, such as suppositories or retention enemas (e.g.,containing conventional suppository bases). In addition to theformulations described previously, the compounds also can be formulatedas a depot preparation. Such long-acting formulations can beadministered by implantation (e.g., subcutaneously or intramuscularly)or by intramuscular injection. Thus, for example, the compounds can beformulated with suitable polymeric or hydrophobic materials (forexample, as an emulsion in an acceptable oil) or ion exchange resins, oras sparingly soluble derivatives, for example, as a sparingly solublesalt.

In particular, a compound disclosed herein can be administered orally,buccally, or sublingually in the form of tablets containing excipients,such as starch or lactose, or in capsules or ovules, either alone or inadmixture with excipients, or in the form of elixirs or suspensionscontaining flavoring or coloring agents. Such liquid preparations can beprepared with pharmaceutically acceptable additives, such as suspendingagents. A compound also can be injected parenterally, for example,intravenously, intramuscularly, subcutaneously, or intracoronarily. Forparenteral administration, the compound is best used in the form of asterile aqueous solution which can contain other substances, forexample, salts, or sugar alcohols, such as mannitol, or glucose, to makethe solution isotonic with blood.

For veterinary use, a compound disclosed herein is administered as asuitably acceptable formulation in accordance with normal veterinarypractice. The veterinarian can readily determine the dosing regimen androute of administration that is most appropriate for a particularanimal.

In some embodiments, all the necessary components for the treatment ofKRAS-related disorder using a compound as disclosed herein either aloneor in combination with another agent or intervention traditionally usedfor the treatment of such disease may be packaged into a kit.Specifically, the present invention provides a kit for use in thetherapeutic intervention of the disease comprising a packaged set ofmedicaments that include the compound disclosed herein as well asbuffers and other components for preparing deliverable forms of saidmedicaments, and/or devices for delivering such medicaments, and/or anyagents that are used in combination therapy with the compound disclosedherein, and/or instructions for the treatment of the disease packagedwith the medicaments. The instructions may be fixed in any tangiblemedium, such as printed paper, or a computer readable magnetic oroptical medium, or instructions to reference a remote computer datasource such as a world wide web page accessible via the interne.

A “therapeutically effective amount” means an amount effective to treator to prevent development of, or to alleviate the existing symptoms of,the subject being treated. Determination of the effective amounts iswell within the capability of those skilled in the art, especially inlight of the detailed disclosure provided herein. Generally, a“therapeutically effective dose” refers to that amount of the compoundthat results in achieving the desired effect. For example, in onepreferred embodiment, a therapeutically effective amount of a compounddisclosed herein decreases KRAS activity by at least 5%, compared tocontrol, at least 10%, at least 15%, at least 20%, at least 25%, atleast 30%, at least 35%, at least 40%, at least 45%, at least 50%, atleast 55%, at least 60%, at least 65%, at least 70%, at least 75%, atleast 80%, at least 85%, or at least 90%.

The amount of compound administered can be dependent on the subjectbeing treated, on the subject's age, health, sex, and weight, the kindof concurrent treatment (if any), severity of the affliction, the natureof the effect desired, the manner and frequency of treatment, and thejudgment of the prescribing physician. The frequency of dosing also canbe dependent on pharmacodynamic effects on arterial oxygen pressures.However, the most preferred dosage can be tailored to the individualsubject, as is understood and determinable by one of skill in the art,without undue experimentation. This typically involves adjustment of astandard dose (e.g., reduction of the dose if the patient has a low bodyweight).

While individual needs vary, determination of optimal ranges ofeffective amounts of the compound is within the skill of the art. Foradministration to a human in the curative or prophylactic treatment ofthe conditions and disorders identified herein, for example, typicaldosages of the compounds of the present invention can be about 0.05mg/kg/day to about 50 mg/kg/day, for example at least 0.05 mg/kg, atleast 0.08 mg/kg, at least 0.1 mg/kg, at least 0.2 mg/kg, at least 0.3mg/kg, at least 0.4 mg/kg, or at least 0.5 mg/kg, and preferably 50mg/kg or less, 40 mg/kg or less, 30 mg/kg or less, 20 mg/kg or less, or10 mg/kg or less, which can be about 2.5 mg/day (0.5 mg/kg×5 kg) toabout 5000 mg/day (50mg/kg×100 kg), for example. For example, dosages ofthe compounds can be about 0.1 mg/kg/day to about 50 mg/kg/day, about0.05 mg/kg/day to about 10 mg/kg/day, about 0.05 mg/kg/day to about 5mg/kg/day, about 0.05 mg/kg/day to about 3 mg/kg/day, about 0.07mg/kg/day to about 3 mg/kg/day, about 0.09 mg/kg/day to about 3mg/kg/day, about 0.05 mg/kg/day to about 0.1 mg/kg/day, about 0.1mg/kg/day to about 1 mg/kg/day, about 1 mg/kg/day to about 10 mg/kg/day,about 1 mg/kg/day to about 5 mg/kg/day, about 1 mg/kg/day to about 3mg/kg/day, about 3 mg/day to about 500 mg/day, about 5 mg/day to about250 mg/day, about 10 mg/day to about 100 mg/day, about 3 mg/day to about10 mg/day, or about 100 mg/day to about 250 mg/day. Such doses may beadministered in a single dose or it may be divided into multiple doses.

Methods of Using KRAS G12C Inhibitors

The present disclosure provides a method of inhibiting RAS-mediated cellsignaling comprising contacting a cell with an effective amount of oneor more compounds disclosed herein. Inhibition of RAS-mediated signaltransduction can be assessed and demonstrated by a wide variety of waysknown in the art. Non-limiting examples include a showing of (a) adecrease in GTPase activity of RAS; (b) a decrease in GTP bindingaffinity or an increase in GDP binding affinity; (c) an increase in Koff of GTP or a decrease in K off of GDP; (d) a decrease in the levelsof signaling transduction molecules downstream in the RAS pathway, suchas a decrease in pMEK, pERK, or pAKT levels; and/or (e) a decrease inbinding of RAS complex to downstream signaling molecules including butnot limited to Raf. Kits and commercially available assays can beutilized for determining one or more of the above.

The disclosure also provides methods of using the compounds orpharmaceutical compositions of the present disclosure to treat diseaseconditions, including but not limited to conditions implicated by G12CKRAS, HRAS or NRAS mutation (e.g., cancer).

In some embodiments, a method for treatment of cancer is provided, themethod comprising administering an effective amount of any of theforegoing pharmaceutical compositions comprising a compound as disclosedherein to a subject in need thereof. In some embodiments, the cancer ismediated by a KRAS, HRAS or NRAS G12C mutation. In various embodiments,the cancer is pancreatic cancer, colorectal cancer or lung cancer. Insome embodiments, the cancer is gall bladder cancer, thyroid cancer, andbile duct cancer.

In some embodiments the disclosure provides method of treating adisorder in a subject in need thereof, wherein the said method comprisesdetermining if the subject has a KRAS, HRAS or NRAS G12C mutation and ifthe subject is determined to have the KRAS, HRAS or NRAS G12C mutation,then administering to the subject a therapeutically effective dose of atleast one compound as disclosed herein or a pharmaceutically acceptablesalt thereof.

The disclosed compounds inhibit anchorage-independent cell growth andtherefore have the potential to inhibit tumor metastasis. Accordingly,another embodiment the disclosure provides a method for inhibiting tumormetastasis, the method comprising administering an effective amount acompound disclosed herein.

KRAS, HRAS or NRAS G12C mutations have also been identified inhematological malignancies (e.g., cancers that affect blood, bone marrowand/or lymph nodes). Accordingly, certain embodiments are directed toadministration of a disclosed compounds (e.g., in the form of apharmaceutical composition) to a patient in need of treatment of ahematological malignancy. Such malignancies include, but are not limitedto leukemias and lymphomas. For example, the presently disclosedcompounds can be used for treatment of diseases such as Acutelymphoblastic leukemia (ALL), Acute myelogenous leukemia (AML), Chroniclymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), Chronicmyelogenous leukemia (CML), Acute monocytic leukemia (AMoL) and/ orother leukemias. In other embodiments, the compounds are useful fortreatment of lymphomas such as all subtypes of Hodgkins lymphoma ornon-Hodgkins lymphoma. In various embodiments, the compounds are usefulfor treatment of plasma cell malignancies such as multiple myeloma,mantle cell lymphoma, and Waldenstrom's macroglubunemia.

Determining whether a tumor or cancer comprises a G12C KRAS, HRAS orNRAS mutation can be undertaken by assessing the nucleotide sequenceencoding the KRAS, HRAS or NRAS protein, by assessing the amino acidsequence of the KRAS, HRAS or NRAS protein, or by assessing thecharacteristics of a putative KRAS, HRAS or NRAS mutant protein. Thesequence of wild-type human KRAS, HRAS or NRAS is known in the art,(e.g. Accession No. NP203524).

Methods for detecting a mutation in a KRAS, HRAS or NRAS nucleotidesequence are known by those of skill in the art. These methods include,but are not limited to, polymerase chain reaction-restriction fragmentlength polymorphism (PCR-RFLP) assays, polymerase chain reaction-singlestrand conformation polymorphism (PCR-SSCP) assays, real-time PCRassays, PCR sequencing, mutant allele-specific PCR amplification (MASA)assays, direct sequencing, primer extension reactions, electrophoresis,oligonucleotide ligation assays, hybridization assays, TaqMan assays,SNP genotyping assays, high resolution melting assays and microarrayanalyses. In some embodiments, samples are evaluated for G12C KRAS, HRASor NRAS mutations by real-time PCR. In real-time PCR, fluorescent probesspecific for the KRAS, HRAS or NRAS G12C mutation are used. When amutation is present, the probe binds and fluorescence is detected. Insome embodiments, the KRAS, HRAS or NRAS G12C mutation is identifiedusing a direct sequencing method of specific regions (e.g., exon 2and/or exon 3) in the KRAS, HRAS or NRAS gene. This technique willidentify all possible mutations in the region sequenced.

Methods for detecting a mutation in a KRAS, HRAS or NRAS protein areknown by those of skill in the art. These methods include, but are notlimited to, detection of a KRAS, HRAS or NRAS mutant using a bindingagent (e.g., an antibody) specific for the mutant protein, proteinelectrophoresis and Western blotting, and direct peptide sequencing.

Methods for determining whether a tumor or cancer comprises a G12C KRAS,HRAS or NRAS mutation can use a variety of samples. In some embodiments,the sample is taken from a subject having a tumor or cancer. In someembodiments, the sample is a fresh tumor/cancer sample. In someembodiments, the sample is a frozen tumor/cancer sample. In someembodiments, the sample is a formalin-fixed paraffin-embedded sample. Insome embodiments, the sample is a circulating tumor cell (CTC) sample.In some embodiments, the sample is processed to a cell lysate. In someembodiments, the sample is processed to DNA or RNA.

The disclosure also relates to a method of treating a hyperproliferativedisorder in a mammal that comprises administering to said mammal atherapeutically effective amount of a compound as disclosed herein, or apharmaceutically acceptable salt thereof. In some embodiments, saidmethod relates to the treatment of a subject who suffers from a cancersuch as acute myeloid leukemia, cancer in adolescents, adrenocorticalcarcinoma childhood, AIDS-related cancers (e.g. Lymphoma and Kaposi'sSarcoma), anal cancer, appendix cancer, astrocytomas, atypical teratoid,basal cell carcinoma, bile duct cancer, bladder cancer, bone cancer,brain stem glioma, brain tumor, breast cancer, bronchial tumors, Burkittlymphoma, carcinoid tumor, atypical teratoid, embryonal tumors, germcell tumor, primary lymphoma, cervical cancer, childhood cancers,chordoma, cardiac tumors, chronic lymphocytic leukemia (CLL), chronicmyelogenous leukemia (CML), chronic myleoproliferative disorders, coloncancer, colorectal cancer, craniopharyngioma, cutaneous T-cell lymphoma,extrahepatic ductal carcinoma in situ (DCIS), embryonal tumors, CNScancer, endometrial cancer, ependymoma, esophageal cancer,esthesioneuroblastoma, ewing sarcoma, extracranial germ cell tumor,extragonadal germ cell tumor, eye cancer, fibrous histiocytoma of bone,gall bladder cancer, gastric cancer, gastrointestinal carcinoid tumor,gastrointestinal stromal tumors (GIST), germ cell tumor, gestationaltrophoblastic tumor, hairy cell leukemia, head and neck cancer, heartcancer, liver cancer, Hodgkin lymphoma, hypopharyngeal cancer,intraocular melanoma, islet cell tumors, pancreatic neuroendocrinetumors, kidney cancer, laryngeal cancer, lip and oral cavity cancer,liver cancer, lobular carcinoma in situ (LCIS), lung cancer, lymphoma,metastatic squamous neck cancer with occult primary, midline tractcarcinoma, mouth cancer, multiple endocrine neoplasia syndromes,multiple myeloma/plasma cell neoplasm, mycosis fungoides,myelodysplastic syndromes,myelodysplastic/myeloproliferative neoplasms,multiple myeloma, merkel cell carcinoma, malignant mesothelioma,malignant fibrous histiocytoma of bone and osteosarcoma, nasal cavityand paranasal sinus cancer, nasopharyngeal cancer, neuroblastoma,non-hodgkin lymphoma, non-small cell lung cancer (NSCLC), oral cancer,lip and oral cavity cancer, oropharyngeal cancer, ovarian cancer,pancreatic cancer, papillomatosis, paraganglioma, paranasal sinus andnasal cavity cancer, parathyroid cancer, penile cancer, pharyngealcancer, pleuropulmonary blastoma, primary central nervous system (CNS)lymphoma, prostate cancer, rectal cancer, transitional cell cancer,retinoblastoma, rhabdomyosarcoma, salivary gland cancer, skin cancer,stomach (gastric) cancer, small cell lung cancer, small intestinecancer, soft tissue sarcoma, T-Cell lymphoma, testicular cancer, throatcancer, thymoma and thymic carcinoma, thyroid cancer, transitional cellcancer of the renal pelvis and ureter, trophoblastic tumor, unusualcancers of childhood, urethral cancer, uterine sarcoma, vaginal cancer,vulvar cancer, or viral-induced cancer. In some embodiments, said methodrelates to the treatment of a non-cancerous hyperproliferative disordersuch as benign hyperplasia of the skin (e.g., psoriasis), restenosis, orprostate (e.g., benign prostatic hypertrophy (BPH)).

In some embodiments, the methods for treatment are directed to treatinglung cancers, the methods comprise administering an effective amount ofany of the above described compound (or a pharmaceutical compositioncomprising the same) to a subject in need thereof. In certainembodiments the lung cancer is a non-small cell lung carcinoma (NSCLC),for example adenocarcinoma, squamous-cell lung carcinoma or large-celllung carcinoma. In some embodiments, the lung cancer is a small celllung carcinoma. Other lung cancers treatable with the disclosedcompounds include, but are not limited to, glandular tumors, carcinoidtumors and undifferentiated carcinomas.

The disclosure further provides methods of modulating a G12C MutantKRAS, HRAS or NRAS protein activity by contacting the protein with aneffective amount of a compound of the disclosure. Modulation can beinhibiting or activating protein activity. In some embodiments, thedisclosure provides methods of inhibiting protein activity by contactingthe G12C Mutant KRAS, HRAS or NRAS protein with an effective amount of acompound of the disclosure in solution. In some embodiments, thedisclosure provides methods of inhibiting the G12C Mutant KRAS, HRAS orNRAS protein activity by contacting a cell, tissue, or organ thatexpresses the protein of interest. In some embodiments, the disclosureprovides methods of inhibiting protein activity in subject including butnot limited to rodents and mammal (e.g., human) by administering intothe subject an effective amount of a compound of the disclosure. In someembodiments, the percentage modulation exceeds 25%, 30%, 40%, 50%, 60%,70%, 80%, or 90%. In some embodiments, the percentage of inhibitingexceeds 25%, 30%, 40%, 50%, 60%, 70%, 80%, or 90%.

In some embodiments, the disclosure provides methods of inhibiting KRAS,HRAS or NRAS G12C activity in a cell by contacting said cell with anamount of a compound of the disclosure sufficient to inhibit theactivity of KRAS, HRAS or NRAS G12C in said cell. In some embodiments,the disclosure provides methods of inhibiting KRAS, HRAS or NRAS G12Cactivity in a tissue by contacting said tissue with an amount of acompound of the disclosure sufficient to inhibit the activity of KRAS,HRAS or NRAS G12C in said tissue. In some embodiments, the disclosureprovides methods of inhibiting KRAS, HRAS or NRAS G12C activity in anorganism by contacting said organism with an amount of a compound of thedisclosure sufficient to inhibit the activity of KRAS, HRAS or NRAS G12Cin said organism. In some embodiments, the disclosure provides methodsof inhibiting KRAS, HRAS or NRAS G12C activity in an animal bycontacting said animal with an amount of a compound of the disclosuresufficient to inhibit the activity of KRAS, HRAS or NRAS G12C in saidanimal. In some embodiments, the disclosure provides methods ofinhibiting KRAS, HRAS or NRAS G12C activity in a mammal by contactingsaid mammal with an amount of a compound of the disclosure sufficient toinhibit the activity of KRAS, HRAS or NRAS G12C in said mammal. In someembodiments, the disclosure provides methods of inhibiting KRAS, HRAS orNRAS G12C activity in a human by contacting said human with an amount ofa compound of the disclosure sufficient to inhibit the activity of KRAS,HRAS or NRAS G12C in said human. The present disclosure provides methodsof treating a disease mediated by KRAS, HRAS or NRAS G12C activity in asubject in need of such treatment.

Combination Therapy

The present disclosure also provides methods for combination therapiesin which an agent known to modulate other pathways, or other componentsof the same pathway, or even overlapping sets of target enzymes are usedin combination with a compound of the present disclosure, or apharmaceutically acceptable salt thereof. In one aspect, such therapyincludes but is not limited to the combination of one or more compoundsof the disclosure with chemotherapeutic agents, therapeutic antibodies,and radiation treatment, to provide a synergistic or additivetherapeutic effect.

Many chemotherapeutics are presently known in the art and can be used incombination with the compounds of the disclosure. In some embodiments,the chemotherapeutic is selected from the group consisting of mitoticinhibitors, alkylating agents, anti-metabolites, intercalatingantibiotics, growth factor inhibitors, cell cycle inhibitors, enzymes,topoisomerase inhibitors, biological response modifiers, anti-hormones,angiogenesis inhibitors, and anti-androgens. Non-limiting examples arechemotherapeutic agents, cytotoxic agents, and non-peptide smallmolecules such as Gleevec® (Imatinib Mesylate), Kyprolis® (carfilzomib),Velcade® (bortezomib), Casodex (bicalutamide), Iressa® (gefitinib),Venclexta™ (venetoclax) and Adriamycin™, (docorubicin) as well as a hostof chemotherapeutic agents. Non-limiting examples of chemotherapeuticagents include alkylating agents such as thiotepa and cyclosphosphamide(Cytoxan™); alkyl sulfonates such as busulfan, improsulfan andpiposulfan; aziridines such as benzodopa, carboquone, meturedopa, anduredopa; ethylenimines and methylamelamines including altretamine,triethylenemelamine, trietylenephosphoramide,triethylenethiophosphaoramide and trimethylolomelamine; nitrogenmustards such as chlorambucil, chlornaphazine, chlorocyclophosphamide,estramustine, ifosfamide, mechlorethamine, mechlorethamine oxidehydrochloride, melphalan, novembichin, phenesterine, prednimustine,trofosfamide, uracil mustard; nitrosureas such as carmustine,chlorozotocin, fotemustine, lomustine, nimustine, ranimustine;antibiotics such as aclacinomysins, actinomycin, authramycin, azaserine,bleomycins, cactinomycin, calicheamicin, carabicin, carminomycin,carzinophilin, Casodex™, chromomycins, dactinomycin, daunorubicin,detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin, epirubicin,esorubicin, idarubicin, marcellomycin, mitomycins, mycophenolic acid,nogalamycin, olivomycins, peplomycin, potfiromycin, puromycin,quelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin,ubenimex, zinostatin, zorubicin; anti-metabolites such as methotrexateand 5-fluorouracil (5-FU); folic acid analogues such as denopterin,methotrexate, pteropterin, trimetrexate; purine analogs such asfludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidineanalogs such as ancitabine, azacitidine, 6-azauridine, carmofur,cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine,androgens such as calusterone, dromostanolone propionate, epitiostanol,mepitiostane, testolactone; anti-adrenals such as aminoglutethimide,mitotane, trilostane; folic acid replenisher such as frolinic acid;aceglatone; aldophosphamide glycoside; aminolevulinic acid; amsacrine;bestrabucil; bisantrene; edatraxate; defofamine; demecolcine;diaziquone; elfomithine; elliptinium acetate; etoglucid; galliumnitrate; hydroxyurea; lentinan; lonidamine; mitoguazone; mitoxantrone;mopidamol; nitracrine; pentostatin; phenamet; pirarubicin; podophyllinicacid; 2-ethylhydrazide; procarbazine; PSK; razoxane; sizofiran;spirogermanium; tenuazonic acid; triaziquone;2,2′,2″-trichlorotriethylamine; urethan; vindesine; dacarbazine;mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine;arabinoside (“Ara-C”); cyclophosphamide; thiotepa; taxanes, e.g.paclitaxel and docetaxel; retinoic acid; esperamicins; capecitabine; andpharmaceutically acceptable salts, acids or derivatives of any of theabove.

Also included as suitable chemotherapeutic cell conditioners areanti-hormonal agents that act to regulate or inhibit hormone action ontumors such as anti-estrogens including for example tamoxifen,(Nolvadex™), raloxifene, aromatase inhibiting 4(5)-imidazoles,4-hydroxytamoxifen, trioxifene, keoxifene, LY 117018, onapristone, andtoremifene (Fareston); and anti-androgens such as flutamide, nilutamide,bicalutamide, leuprolide, and goserelin; chlorambucil; gemcitabine;6-thioguanine; mercaptopurine; methotrexate; platinum analogs such ascisplatin and carboplatin; vinblastine; platinum; etoposide (VP-16);ifosfamide; mitomycin C; mitoxantrone; vincristine; vinorelbine;navelbine; novantrone; teniposide; daunomycin; aminopterin; xeloda;ibandronate; camptothecin-11 (CPT-11); topoisomerase inhibitor RFS 2000;difluoromethylornithine (DMFO).

Where desired, the compounds or pharmaceutical composition of thepresent disclosure can be used in combination with commonly prescribedanti-cancer drugs such as Herceptin®, Avastin®, Erbitux®, Rituxan®,Taxol®, Arimidex®, Taxotere®, ABVD, AVICINE, Abagovomab, Acridinecarboxamide, Adecatumumab, 17-N-Allylamino-17-demethoxygeldanamycin,Alpharadin, Alvocidib, 3-Aminopyridine-2-carboxaldehydethiosemicarbazone, Amonafide, Anthracenedione, Anti-CD22 immunotoxins,Antineoplastic, Antitumorigenic herbs, Apaziquone, Atiprimod,Azathioprine, Belotecan, Bendamustine, BIBW 2992, Biricodar,Brostallicin, Bryostatin, Buthionine sulfoximine, CBV (chemotherapy),Calyculin, cell-cycle nonspecific antineoplastic agents, Dichloroaceticacid, Discodermolide, Elsamitrucin, Enocitabine, Epothilone, Eribulin,Everolimus, Exatecan, Exisulind, Ferruginol, Forodesine, Fosfestrol, ICEchemotherapy regimen, IT-101, Imexon, Imiquimod, Indolocarbazole,Irofulven, Laniquidar, Larotaxel, Lenalidomide, Lucanthone, Lurtotecan,Mafosfamide, Mitozolomide, Nafoxidine, Nedaplatin, Olaparib, Ortataxel,PAC-1, Pawpaw, Pixantrone, Proteasome inhibitor, Rebeccamycin,Resiquimod, Rubitecan, SN-38, Salinosporamide A, Sapacitabine, StanfordV, Swainsonine, Talaporfin, Tariquidar, Tegafur-uracil, Temodar,Tesetaxel, Triplatin tetranitrate, Tris(2-chloroethyl)amine,Troxacitabine, Uramustine, Vadimezan, Vinflunine, ZD6126 or Zosuquidar.

This disclosure further relates to a method for using the compounds orpharmaceutical compositions provided herein, in combination withradiation therapy for inhibiting abnormal cell growth or treating thehyperproliferative disorder in the mammal. Techniques for administeringradiation therapy are known in the art, and these techniques can be usedin the combination therapy described herein. The administration of thecompound of the disclosure in this combination therapy can be determinedas described herein.

Radiation therapy can be administered through one of several methods, ora combination of methods, including without limitation external-beamtherapy, internal radiation therapy, implant radiation, stereotacticradiosurgery, systemic radiation therapy, radiotherapy and permanent ortemporary interstitial brachytherapy. The term “brachytherapy,” as usedherein, refers to radiation therapy delivered by a spatially confinedradioactive material inserted into the body at or near a tumor or otherproliferative tissue disease site. The term is intended withoutlimitation to include exposure to radioactive isotopes (e.g. At-211,1-131, 1-125, Y-90, Re-186, Re-188, Sm-153, Bi-212, P-32, andradioactive isotopes of Lu). Suitable radiation sources for use as acell conditioner of the present disclosure include both solids andliquids. By way of non-limiting example, the radiation source can be aradionuclide, such as 1-125, I-131, Yb-169, Ir-192 as a solid source,1-125 as a solid source, or other radionuclides that emit photons, betaparticles, gamma radiation, or other therapeutic rays. The radioactivematerial can also be a fluid made from any solution of radionuclide(s),e.g., a solution of 1-125 or 1-131, or a radioactive fluid can beproduced using a slurry of a suitable fluid containing small particlesof solid radionuclides, such as Au-198, Y-90. Moreover, theradionuclide(s) can be embodied in a gel or radioactive micro spheres.

The compounds or pharmaceutical compositions of the disclosure can beused in combination with an amount of one or more substances selectedfrom anti-angiogenesis agents, signal transduction inhibitors,antiproliferative agents, glycolysis inhibitors, or autophagyinhibitors.

Anti-angiogenesis agents, such as MMP-2 (matrix-metalloproteinase 2)inhibitors, MMP-9 (matrix-metalloproteinase 9) inhibitors, and COX-11(cyclooxygenase 11) inhibitors, can be used in conjunction with acompound of the disclosure and pharmaceutical compositions describedherein. Anti-angiogenesis agents include, for example, rapamycin,temsirolimus (CCI-779), everolimus (RAD001), sorafenib, sunitinib, andbevacizumab. Examples of useful COX-II inhibitors include alecoxib,valdecoxib, and rofecoxib. Examples of useful matrix metalloproteinaseinhibitors are described in WO 96/33172 WO 96/27583 European PatentPublication EP0818442, European Patent Publication EP1004578, WO98/07697, WO 98/03516, WO 98/34918, WO 98/34915, WO 98/33768, WO98/30566, European Patent Publication 606046, European PatentPublication 931 788, WO 90/05719, WO 99/52910, WO 99/52889, WO 99/29667,WO1999007675, European Patent Publication EP1786785, European PatentPublication No. EP1181017, United States Publication No. US20090012085,United States Publication U.S. Pat. Nos. 5,863,949, 5,861,510, andEuropean Patent Publication EP0780386, all of which are incorporatedherein in their entireties by reference. Preferred MMP-2 and MMP-9inhibitors are those that have little or no activity inhibiting MMP-1.More preferred, are those that selectively inhibit MMP-2 and/or AMP-9relative to the other matrix-metalloproteinases (i.e., MAP-1, MMP-3,MMP-4, MMP-5, MMP-6, MMP-7, MMP-8, MMP-10, MMP-11, MMP-12, andMMP-13).Some specific examples of MMP inhibitors useful in the disclosure areAG-3340, RO 32-3555, and RS 13-0830.

The present compounds may also be used in co-therapies with otheranti-neoplastic agents, such as acemannan, aclarubicin, aldesleukin,alemtuzumab, alitretinoin, altretamine, amifostine, aminolevulinic acid,amrubicin, amsacrine, anagrelide, anastrozole, ANCER, ancestim,ARGLABIN, arsenic trioxide, BAM 002 (Novelos), bexarotene, bicalutamide,broxuridine, capecitabine, celmoleukin, cetrorelix, cladribine,clotrimazole, cytarabine ocfosfate, DA 3030 (Dong-A), daclizumab,denileukin diftitox, deslorelin, dexrazoxane, dilazep, docetaxel,docosanol, doxercalciferol, doxifluridine, doxorubicin, bromocriptine,carmustine, cytarabine, fluorouracil, HIT diclofenac, interferon alfa,daunorubicin, doxorubicin, tretinoin, edelfosine, edrecolomab,eflornithine, emitefur, epirubicin, epoetin beta, etoposide phosphate,exemestane, exisulind, fadrozole, filgrastim, finasteride, fludarabinephosphate, formestane, fotemustine, gallium nitrate, gemcitabine,gemtuzumab zogamicin, gimeracil/oteracil/tegafur combination, glycopine,goserelin, heptaplatin, human chorionic gonadotropin, human fetal alphafetoprotein, ibandronic acid, idarubicin, (imiquimod, interferon alfa,interferon alfa, natural, interferon alfa-2, interferon alfa-2a,interferon alfa-2b, interferon alfa-N1, interferon alfa-_(n3),interferon alfacon-1, interferon alpha, natural, interferon beta,interferon beta-1a, interferon beta-1b, interferon gamma, naturalinterferon gamma-1a, interferon gamma-1b, interleukin-1 beta,iobenguane, irinotecan, irsogladine, lanreotide, LC 9018 (Yakult),leflunomide, lenograstim, lentinan sulfate, letrozole, leukocyte alphainterferon, leuprorelin, levamisole+fluorouracil, liarozole, lobaplatin,lonidamine, lovastatin, masoprocol, melarsoprol, metoclopramide,mifepristone, miltefosine, mirimostim, mismatched double stranded RNA,mitoguazone, mitolactol, mitoxantrone, molgramostim, nafarelin,naloxone+pentazocine, nartograstim, nedaplatin, nilutamide, noscapine,novel erythropoiesis stimulating protein, NSC 631570 octreotide,oprelvekin, osaterone, oxaliplatin, paclitaxel, pamidronic acid,pegaspargase, peginterferon alfa-2b, pentosan polysulfate sodium,pentostatin, picibanil, pirarubicin, rabbit antithymocyte polyclonalantibody, polyethylene glycol interferon alfa-2a, porfimer sodium,raloxifene, raltitrexed, rasburiembodiment, rhenium Re 186 etidronate,RH retinamide, rituximab, romurtide, samarium (153 Sm) lexidronam,sargramostim, sizofiran, sobuzoxane, sonermin, strontium-89 chloride,suramin, tasonermin, tazarotene, tegafur, temoporfin, temozolomide,teniposide, tetrachlorodecaoxide, thalidomide, thymalfasin, thyrotropinalfa, topotecan, toremifene, tositumomab-iodine 131, trastuzumab,treosulfan, tretinoin, trilostane, trimetrexate, triptorelin, tumornecrosis factor alpha, natural, ubenimex, bladder cancer vaccine,Maruyama vaccine, melanoma lysate vaccine, valrubicin, verteporfin,vinorelbine, VIRULIZIN, zinostatin stimalamer, or zoledronic acid;abarelix; AE 941 (Aeterna), ambamustine, antisense oligonucleotide,bcl-2 (Genta), APC 8015 (Dendreon), cetuximab, decitabine,dexaminoglutethimide, diaziquone, EL 532 (Elan), EM 800 (Endorecherche),eniluracil, etanidazole, fenretinide, filgrastim SD01 (Amgen),fulvestrant, galocitabine, gastrin 17 immunogen, HLA-B7 gene therapy(Vical), granulocyte macrophage colony stimulating factor, histaminedihydrochloride, ibritumomab tiuxetan, ilomastat, IM 862 (Cytran),interleukin-2, iproxifene, LDI 200 (Milkhaus), leridistim, lintuzumab,CA 125 MAb (Biomira), cancer MAb (Japan Pharmaceutical Development),HER-2 and Fc MAb (Medarex), idiotypic 105AD7 MAb (CRC Technology),idiotypic CEA MAb (Trilex), LYM-1-iodine 131 MAb (Techniclone),polymorphic epithelial mucin-yttrium 90 MAb (Antisoma), marimastat,menogaril, mitumomab, motexafin gadolinium, MX 6 (Galderma), nelarabine,nolatrexed, P 30 protein, pegvisomant, pemetrexed, porfiromycin,prinomastat, RL 0903 (Shire), rubitecan, satraplatin, sodiumphenylacetate, sparfosic acid, SRL 172 (SR Pharma), SU 5416 (SUGEN, nowPfizer, Inc.), TA 077 (Tanabe), tetrathiomolybdate, thaliblastine,thrombopoietin, tin ethyl etiopurpurin, tirapazamine, cancer vaccine(Biomira), melanoma vaccine (New York University), melanoma vaccine(Sloan Kettering Institute), melanoma oncolysate vaccine (New YorkMedical College), viral melanoma cell lysates vaccine (Royal NewcastleHospital), or valspodar.

The compounds of the invention may further be used with VEGFRinhibitors. Other compounds described in the following patents andpatent applications can be used in combination therapy: U.S. Pat. No.6,258,812, US 2003/0105091, WO 01/37820, U.S. Pat. No. 6,235,764, WO01/32651, U.S. Pat. Nos. 6,630,500, 6,515,004, 6,713,485, 5,521,184,5,770,599, 5,747,498, WO 02/68406, WO 02/66470, WO 02/55501, WO04/05279, WO 04/07481, WO 04/07458, WO 04/09784, WO 02/59110, WO99/45009, WO 00/59509, WO 99/61422, U.S. Pat. No. 5,990,141, WO00/12089, and WO 00/02871.

In some embodiments, the combination comprises a composition of thepresent invention in combination with at least one anti-angiogenicagent. Agents are inclusive of, but not limited to, in vitrosynthetically prepared chemical compositions, antibodies, antigenbinding regions, radionuclides, and combinations and conjugates thereof.An agent can be an agonist, antagonist, allosteric modulator, toxin or,more generally, may act to inhibit or stimulate its target (e.g.,receptor or enzyme activation or inhibition), and thereby promote celldeath or arrest cell growth.

Exemplary anti-angiogenic agents include ERBITUX™ (IMC-C225), KDR(kinase domain receptor) inhibitory agents (e.g., antibodies and antigenbinding regions that specifically bind to the kinase domain receptor),anti-VEGF agents (e.g., antibodies or antigen binding regions thatspecifically bind VEGF, or soluble VEGF receptors or a ligand bindingregion thereof) such as AVASTIN™ or VEGF-TRAP™, and anti-VEGF receptoragents (e.g., antibodies or antigen binding regions that specificallybind thereto), EGFR inhibitory agents (e.g., antibodies or antigenbinding regions that specifically bind thereto) such as Vectibix(panitumumab), IRESSA™ (gefitinib), TARCEVA™ (erlotinib), anti-Ang1 andanti-Ang2 agents (e.g., antibodies or antigen binding regionsspecifically binding thereto or to their receptors, e.g., Tie2/Tek), andanti-Tie2 kinase inhibitory agents (e.g., antibodies or antigen bindingregions that specifically bind thereto). The pharmaceutical compositionsof the present invention can also include one or more agents (e.g.,antibodies, antigen binding regions, or soluble receptors) thatspecifically bind and inhibit the activity of growth factors, such asantagonists of hepatocyte growth factor (HGF, also known as ScatterFactor), and antibodies or antigen binding regions that specificallybind its receptor “c-met”.

Other anti-angiogenic agents include Campath, IL-8, B-FGF, Tekantagonists (Ceretti et al., U.S. Publication No. 2003/0162712; U.S.Pat. No. 6,413,932), anti-TWEAK agents (e.g., specifically bindingantibodies or antigen binding regions, or soluble TWEAK receptorantagonists; see, Wiley, U.S. Pat. No. 6,727,225), ADAM distintegrindomain to antagonize the binding of integrin to its ligands (Fanslow etal., U.S. Publication No. 2002/0042368), specifically binding anti-ephreceptor and/or anti-ephrin antibodies or antigen binding regions (U.S.Pat. Nos. 5,981,245; 5,728,813; 5,969,110; 6,596,852; 6,232,447;6,057,124 and patent family members thereof), and anti-PDGF-BBantagonists (e.g., specifically binding antibodies or antigen bindingregions) as well as antibodies or antigen binding regions specificallybinding to PDGF-BB ligands, and PDGFR kinase inhibitory agents (e.g.,antibodies or antigen binding regions that specifically bind thereto).Additional anti-angiogenic/anti-tumor agents include: SD-7784 (Pfizer,USA); cilengitide.(Merck KGaA, Germany, EPO 770622); pegaptaniboctasodium, (Gilead Sciences, USA); Alphastatin, (BioActa, UK); M-PGA,(Celgene, USA, U.S. Pat. No. 5712291); ilomastat, (Arriva, USA, U.S.Pat. No. 5892112); emaxanib, (Pfizer, USA, U.S. Pat. No. 5792783);vatalanib, (Novartis, Switzerland); 2-methoxyestradiol, (EntreMed, nowCASI Pharamaceuticals, USA); TLC ELL-12, (Elan, Ireland); anecortaveacetate, (Alcon, USA); alpha-D148 Mab, (Amgen, USA); CEP-7055,(Cephalon,USA); anti-Vn Mab, (Crucell, Netherlands) DAC: antiangiogenic,(ConjuChem, Canada); Angiocidin, (InKine Pharmaceutical, USA); KM-2550,(Kyowa Hakko, Japan); SU-0879, (Pfizer, USA); CGP-79787, (Novartis,Switzerland, EP 970070); ARGENT technology, (Ariad, USA); YIGSR-Stealth,(Johnson & Johnson, USA); fibrinogen-E fragment, (BioActa, UK);angiogenesis inhibitor, (Trigen, UK); TBC-1635, (EncysivePharmaceuticals, USA); SC-236, (Pfizer, USA); ABT-567, (Abbott, USA);Metastatin, (EntreMed, USA); angiogenesis inhibitor, (Tripep, Sweden);maspin, (Sosei, Japan); 2-methoxyestradiol, (Oncology SciencesCorporation, USA); ER-68203-00, (IVAX, USA); Benefin, (Lane Labs, USA);Tz-93, (Tsumura, Japan); TAN-1120, (Takeda, Japan); FR-111142,(Fujisawa, Japan, JP 02233610); platelet factor 4, (RepliGen, USA, EP407122); vascular endothelial growth factor antagonist, (Borean,Denmark); bevacizumab (pINN), (Genentech, USA); angiogenesis inhibitors,(SUGEN, USA); XL 784, (Exelixis, USA); XL 647, (Exelixis, USA); MAb,alpha5beta3 integrin, second generation, (Applied Molecular Evolution,USA and MedImmune, USA); gene therapy, retinopathy, (Oxford BioMedica,UK); enzastaurin hydrochloride (USAN), (Lilly, USA); CEP 7055,(Cephalon, USA and Sanofi-Synthelabo, France); BC 1, (Genoa Institute ofCancer Research, Italy); angiogenesis inhibitor, (Alchemia, Australia);VEGF antagonist, (Regeneron, USA); rBPI 21 and BPI-derivedantiangiogenic, (XOMA, USA); PI 88, (Progen, Australia); cilengitide(pINN), (Merck KGaA, German; Munich Technical University, Germany,Scripps Clinic and Research Foundation, USA); cetuximab (INN), (Aventis,France); AVE 8062, (Ajinomoto, Japan); AS 1404, (Cancer ResearchLaboratory, New Zealand); SG 292, (Telios, USA); Endostatin, (BostonChildrens Hospital, USA); ATN 161, (Attenuon, USA); ANGIOSTATIN, (BostonChildrens Hospital, USA); 2-methoxyestradiol, (Boston ChildrensHospital, USA); ZD 6474, (AstraZeneca, UK); ZD 6126, (AngiogenePharmaceuticals, UK); PPI 2458, (Praecis, USA); AZD 9935, (AstraZeneca,UK); AZD 2171, (AstraZeneca, UK); vatalanib (pINN), (Novartis,Switzerland and Schering AG, Germany); tissue factor pathway inhibitors,(EntreMed, USA); pegaptanib (Pinn), (Gilead Sciences, USA);xanthorrhizol, (Yonsei University, South Korea); vaccine, gene-based,VEGF-2, (Scripps Clinic and Research Foundation, USA); SPV5.2,(Supratek, Canada); SDX 103, (University of California at San Diego,USA); PX 478, (ProlX, USA); METASTATIN, (EntreMed, now CASIPharmaceuticals, USA); troponin I, (Harvard University, USA); SU 6668,(SUGEN, now Pfizer, Inc., USA); OXI 4503, (OXiGENE, USA); o-guanidines,(Dimensional Pharmaceuticals, USA); motuporamine C, (British ColumbiaUniversity, Canada); CDP 791, (Celltech Group, UK); atiprimod (pINN),(GlaxoSmithKline, UK); E 7820, (Eisai, Japan); CYC 381, (HarvardUniversity, USA); AE 941, (Aeterna, Canada); vaccine, angiogenesis,(EntreMed, now CASI Pharmaceuticals, USA); urokinase plasminogenactivator inhibitor, (Dendreon, USA); oglufanide (pINN), (Melmotte,USA); HIF-lalfa inhibitors, (Xenova, UK); CEP 5214, (Cephalon, USA); BAYRES 2622, (Bayer, Germany); Angiocidin, (InKine, USA); A6, (Angstrom,USA); KR 31372, (Korea Research Institute of Chemical Technology, SouthKorea); GW 2286, (GlaxoSmithKline, UK); EHT 0101, (ExonHit, France); CP868596, (Pfizer, USA); CP 564959, (OSI, USA); CP 547632, (Pfizer, USA);786034, (GlaxoSmithKline, UK); KRN 633, (Kirin Brewery, Japan); drugdelivery system, intraocular, 2-methoxyestradiol, (EntreMed, USA);anginex, (Maastricht University, Netherlands, and Minnesota University,USA); ABT 510, (Abbott, USA); AAL 993, (Novartis, Switzerland); VEGI,(ProteomTech, USA); tumor necrosis factor-alpha inhibitors, (NationalInstitute on Aging, USA); SU 11248, (Pfizer, USA and SUGEN USA); ABT518, (Abbott, USA); YH16, (Yantai Rongchang, China); S-3APG, (BostonChildrens Hospital, USA and EntreMed, USA); MAb, KDR, (ImClone Systems,USA); MAb, alpha5 betal, (Protein Design, USA); KDR kinase inhibitor,(Celltech Group, UK, and Johnson & Johnson, USA); GFB 116, (SouthFlorida University, USA and Yale University, USA); CS 706, (Sankyo,Japan); combretastatin A4 prodrug, (Arizona State University, USA);chondroitinase AC, (IBEX, Canada); BAY RES 2690, (Bayer, Germany); AGM1470, (Harvard University, USA, Takeda, Japan, and TAP, USA); AG 13925,(Agouron, USA); Tetrathiomolybdate, (University of Michigan, USA); GCS100, (Wayne State University, USA) CV 247, (Ivy Medical, UK); CKD 732,(Chong Kun Dang, South Korea); MAb, vascular endothelium growth factor,(Xenova, UK); irsogladine (INN), (Nippon Shinyaku, Japan); RG 13577,(Aventis, France); WX 360, (Wilex, Germany); squalamine (pINN),(Genaera, USA); RPI 4610, (Sirna, USA); cancer therapy, (Marinova,Australia); heparanase inhibitors, (InSight, Israel); KL 3106, (Kolon,South Korea); Honokiol, (Emory University, USA); ZK CDK, (Schering AG,Germany); ZK Angio, (Schering AG, Germany); ZK 229561, (Novartis,Switzerland, and Schering AG, Germany); XMP 300, (XOMA, USA); VGA 1102,(Taisho, Japan); VEGF receptor modulators, (Pharmacopeia, USA);VE-cadherin-2 antagonists, (ImClone Systems, USA); Vasostatin, (NationalInstitutes of Health, USA);vaccine, Flk-1, (ImClone Systems, USA); TZ93, (Tsumura, Japan); TumStatin, (Beth Israel Hospital, USA); truncatedsoluble FLT 1 (vascular endothelial growth factor receptor 1), (Merck &Co, USA); Tie-2 ligands, (Regeneron, USA); and, thrombospondin 1inhibitor, (Allegheny Health, Education and Research Foundation, USA).

Autophagy inhibitors include, but are not limited to chloroquine,3-methyladenine, hydroxychloroquine (Plaquenil™), bafilomycin A1,5-amino-4-imidazole carboxamide riboside (AICAR), okadaic acid,autophagy-suppressive algal toxins which inhibit protein phosphatases oftype 2A or type 1, analogues of cAMP, and drugs which elevate cAMPlevels such as adenosine, LY204002, N6-mercaptopurine riboside, andvinblastine. In addition, antisense or siRNA that inhibits expression ofproteins including but not limited to ATGS (which are implicated inautophagy), may also be used.

Additional pharmaceutically active compounds/agents that can be used inthe treatment of cancers and that can be used in combination with one ormore compound of the present invention include: epoetin alfa;darbepoetin alfa; panitumumab; pegfilgrastim; palifermin; filgrastim;denosumab; ancestim; AMG 102; AMG 176; AMG 386; AMG 479; AMG 655; AMG745; AMG 951; and AMG 706, or a pharmaceutically acceptable saltthereof.

In certain embodiments, a composition provided herein is conjointlyadministered with a chemotherapeutic agent. Suitable chemotherapeuticagents may include, natural products such as vinca alkaloids (e.g.,vinblastine, vincristine, and vinorelbine), paclitaxel,epidipodophyllotoxins (e.g., etoposide and teniposide), antibiotics(e.g., dactinomycin (actinomycin D), daunorubicin, doxorubicin, andidarubicin), anthracyclines, mitoxantrone, bleomycins, plicamycin(mithramycin), mitomycin, enzymes (e.g., L-asparaginase whichsystemically metabolizes L-asparagine and deprives cells which do nothave the capacity to synthesize their own asparagine), antiplateletagents, antiproliferative/antimitotic alkylating agents such as nitrogenmustards (e.g., mechlorethamine, cyclophosphamide and analogs,melphalan, and chlorambucil), ethylenimines and methylmelamines (e.g.,hexaamethylmelaamine and thiotepa), CDK inhibitors (e.g., seliciclib,UCN-01, P1446A-05, PD-0332991, dinaciclib, P27-00, AT-7519, RGB286638,and SCH727965), alkyl sulfonates (e.g., busulfan), nitrosoureas (e.g.,carmustine (BCNU) and analogs, and streptozocin), trazenes-dacarbazinine(DTIC), antiproliferative/antimitotic antimetabolites such as folic acidanalogs (e.g., methotrexate), pyrimidine analogs (e.g., fluorouracil,floxuridine, and cytarabine), purine analogs and related inhibitors(e.g., mercaptopurine, thioguanine, pentostatin and2-chlorodeoxyadenosine), aromatase inhibitors (e.g., anastrozole,exemestane, and letrozole), and platinum coordination complexes (e.g.,cisplatin and carboplatin), procarbazine, hydroxyurea, mitotane,aminoglutethimide, histone deacetylase (HDAC) inhibitors (e.g.,trichostatin, sodium butyrate, apicidan, suberoyl anilide hydroamicacid, vorinostat, LBH 589, romidepsin, ACY-1215, and panobinostat), mTorinhibitors (e.g., temsirolimus, everolimus, ridaforolimus, andsirolimus), KSP(Eg5) inhibitors (e.g., Array 520), DNA binding agents(e.g., Zalypsis), PI3K delta inhibitor (e.g., GS-1101 and TGR-1202),PI3K delta and gamma inhibitor (e.g., CAL-130), multi-kinase inhibitor(e.g., TGO2 and sorafenib), hormones (e.g., estrogen) and hormoneagonists such as leutinizing hormone releasing hormone (LHRH) agonists(e.g., goserelin, leuprolide and triptorelin), BAFF-neutralizingantibody (e.g., LY2127399), IKK inhibitors, p38MAPK inhibitors,anti-IL-6 (e.g., CNT0328), telomerase inhibitors (e.g., GRN 163 L),aurora kinase inhibitors (e.g., MLN8237), cell surface monoclonalantibodies (e.g., anti-CD38 (HUMAX-CD38), anti-CS1 (e.g., elotuzumab),HSP90 inhibitors (e.g., 17 AAG and KOS 953), PI3K/Akt inhibitors (e.g.,perifosine), Akt inhibitor (e.g., GSK-2141795), PKC inhibitors (e.g.,enzastaurin), FTIs (e.g., Zarnestra™), anti-CD138 (e.g., BT062), Torc1/2specific kinase inhibitor (e.g., INK128), kinase inhibitor (e.g.,GS-1101), ER/UPR targeting agent (e.g., MKC-3946), cFMS inhibitor (e.g.,ARRY-382), JAK1/2 inhibitor (e.g., CYT387), PARP inhibitor (e.g.,olaparib and veliparib (ABT-888)), BCL-2 antagonist. Otherchemotherapeutic agents may include mechlorethamine, camptothecin,ifosfamide, tamoxifen, raloxifene, gemcitabine, navelbine, sorafenib, orany analog or derivative variant of the foregoing.

The compounds of the present invention may also be used in combinationwith radiation therapy, hormone therapy, surgery and immunotherapy,which therapies are well known to those skilled in the art.

In certain embodiments, a pharmaceutical composition provided herein isconjointly administered with a steroid. Suitable steroids may include,but are not limited to, 21-acetoxypregnenolone, alclometasone,algestone, amcinonide, beclomethasone, betamethasone, budesonide,chloroprednisone, clobetasol, clocortolone, cloprednol, corticosterone,cortisone, cortivazol, deflazacort, desonide, desoximetasone,dexamethasone, diflorasone, diflucortolone, difuprednate, enoxolone,fluazacort, flucloronide, flumethasone, flunisolide, fluocinoloneacetonide, fluocinonide, fluocortin butyl, fluocortolone,fluorometholone, fluperolone acetate, fluprednidene acetate,fluprednisolone, flurandrenolide, fluticasone propionate, formocortal,halcinonide, halobetasol propionate, halometasone, hydrocortisone,loteprednol etabonate, mazipredone, medrysone, meprednisone,methylprednisolone, mometasone furoate, paramethasone, prednicarbate,prednisolone, prednisolone 25-diethylaminoacetate, prednisolone sodiumphosphate, prednisone, prednival, prednylidene, rimexolone, tixocortol,triamcinolone, triamcinolone acetonide, triamcinolone benetonide,triamcinolone hexacetonide, and salts and/or derivatives thereof. In aparticular embodiment, the compounds of the present invention can alsobe used in combination with additional pharmaceutically active agentsthat treat nausea. Examples of agents that can be used to treat nauseainclude: dronabinol; granisetron; metoclopramide; ondansetron; andprochlorperazine; or a pharmaceutically acceptable salt thereof.

The compounds of the present invention may also be used in combinationwith an additional pharmaceutically active compound that disrupts orinhibits RAS-RAF-ERK or PI3K-AKT-TOR signaling pathways. In other suchcombinations, the additional pharmaceutically active compound is a PD-1and PD-L1 antagonist. The compounds or pharmaceutical compositions ofthe disclosure can also be used in combination with an amount of one ormore substances selected from EGFR inhibitors, MEK inhibitors, PI3Kinhibitors, AKT inhibitors, TOR inhibitors, Mcl-1 inhibitors, BCL-2inhibitors, SHP2 inhibitors, proteasome inhibitors, and immunetherapies, including monoclonal antibodies, immunomodulatory imides(IMiDs), anti-PD-1, anti-PDL-1, anti-CTLA4, anti-LAG1, and anti-OX40agents, GITR agonists, CAR-T cells, and BiTEs.

EGFR inhibitors include, but are not limited to, small moleculeantagonists, antibody inhibitors, or specific antisense nucleotide orsiRNA. Useful antibody inhibitors of EGFR include cetuximab (Erbitux),panitumumab (Vectibix), zalutumumab, nimotuzumab, and matuzumab. Smallmolecule antagonists of EGFR include gefitinib, erlotinib (Tarceva), andmost recently, lapatinib (TykerB). See e.g., Yan L, et. al.,Pharmacogenetics and Pharmacogenomics In Oncology Therapeutic AntibodyDevelopment, BioTechniques 2005; 39(4): 565-8, and Paez J G, et. al.,EGFR Mutations In Lung Cancer Correlation With Clinical Response ToGefitinib Therapy, Science 2004; 304(5676): 1497-500.

Non-limiting examples of small molecule EGFR inhibitors include any ofthe EGFR inhibitors described in the following patent publications, andall pharmaceutically acceptable salts and solvates of said EGFRinhibitors: European Patent Application EP 520722, published Dec. 30,1992; European Patent Application EP 566226, published Oct. 20, 1993;PCT International Publication WO 96/33980, published Oct. 31, 1996; U.S.Pat. No. 5,747,498, issued May 5, 1998; PCT International Publication WO96/30347, published Oct. 3, 1996; European Patent Application EP 787772,published Aug. 6, 1997; PCT International Publication WO 97/30034,published Aug. 21, 1997; PCT International Publication WO 97/30044,published Aug. 21, 1997; PCT International Publication WO 97/38994,published Oct. 23, 1997; PCT International Publication WO 97/49688,published Dec. 31, 1997; European Patent Application EP 837063,published Apr. 22, 1998; PCT International Publication WO 98/02434,published Jan. 22, 1998; PCT International Publication WO 97/38983,published Oct. 23, 1997; PCT International Publication WO 95/19774,published Jul. 27, 1995; PCT International Publication WO 95/19970,published Jul. 27, 1995; PCT International Publication WO 97/13771,published Apr. 17, 1997; PCT International Publication WO 98/02437,published Jan. 22, 1998; PCT International Publication WO 98/02438,published Jan. 22, 1998; PCT International Publication WO 97/32881,published Sep. 12, 1997; German Application DE 19629652, published Jan.29, 1998; PCT International Publication WO 98/33798, published Aug. 6,1998; PCT International Publication WO 97/32880, published Sep. 12,1997; PCT International Publication WO 97/32880 published Sep. 12, 1997;European Patent Application EP 682027, published Nov. 15, 1995; PCTInternational Publication WO 97/02266, published Jan. 23, 197; PCTInternational Publication WO 97/27199, published Jul. 31, 1997; PCTInternational Publication WO 98/07726, published Feb. 26, 1998; PCTInternational Publication WO 97/34895, published Sep. 25, 1997; PCTInternational Publication WO 96/31510′, published Oct. 10, 1996; PCTInternational Publication WO 98/14449, published Apr. 9, 1998; PCTInternational Publication WO 98/14450, published Apr. 9, 1998; PCTInternational Publication WO 98/14451, published Apr. 9, 1998; PCTInternational Publication WO 95/09847, published Apr. 13, 1995; PCTInternational Publication WO 97/19065, published May 29, 1997; PCTInternational Publication WO 98/17662, published Apr. 30, 1998; U.S.Pat. No. 5,789,427, issued Aug. 4, 1998; U.S. Pat. No. 5,650,415, issuedJul. 22, 1997; U.S. Pat. No. 5,656,643, issued Aug. 12, 1997; PCTInternational Publication WO 99/35146, published Jul. 15, 1999; PCTInternational Publication WO 99/35132, published Jul. 15, 1999; PCTInternational Publication WO 99/07701, published Feb. 18, 1999; and PCTInternational Publication WO 92/20642 published Nov. 26, 1992.Additional non-limiting examples of small molecule EGFR inhibitorsinclude any of the EGFR inhibitors described in Traxler, P., 1998, Exp.Opin. Ther. Patents 8(12):1599-1625.

Antibody-based EGFR inhibitors include any anti-EGFR antibody orantibody fragment that can partially or completely block EGFR activationby its natural ligand. Non-limiting examples of antibody-based EGFRinhibitors include those described in Modjtahedi, H., et al., 1993, Br.J. Cancer 67:247-253; Teramoto, T., et al., 1996, Cancer 77:639-645;Goldstein et al., 1995, Clin. Cancer Res. 1:1311-1318; Huang, S. M., etal., 1999, Cancer Res. 15:59(8):1935-40; and Yang, X., et al., 1999,Cancer Res. 59:1236-1243. Thus, the EGFR inhibitor can be monoclonalantibody Mab E7.6.3 (Yang, 1999 supra), or Mab C225 (ATCC Accession No.HB-8508), or an antibody or antibody fragment having the bindingspecificity thereof.

The KRAS^(G12C) inhibitors of the present invention can be used incombination with MEK inhibitors. Particular MEK inhibitors that can beused in the combinations of the present invention include PD-325901,trametinib, pimasertib, MEK162 [also known as binimetinib], TAK-733,GDC-0973 and AZD8330. A particular MEK inhibitor that can be used alongwith KRAS^(G12C) inhibitor in the combinations of the present inventionis trametinib (tradename: Mekinist®, commercially available fromNovartis Pharmaceuticals Corp.). Another particular MEK inhibitor isN-(((2R)-2,3-dihydroxypropyl)oxy)-3,4-difluoro-2-((2-fluoro-4-iodophenyl)amino)benzamide,also known as AMG 1009089, 1009089 or PD-325901.

Another particular MEK inhibitor that can be used in the combinations ofthe present invention includes cobimetinib. MEK inhibitors include, butare not limited to, CI-1040, AZD6244, PD318088, PD98059, PD334581,RDEA119, ARRY-142886, and ARRY-438162.

PI3K inhibitors include, but are not limited to, wortmannin,17-hydroxywortmannin analogs described in WO 06/044453,4-[2-(1H-Indazol-4-yl)-6-[[4-(methylsulfonyl)piperazin-1-yl]methyl]thieno[3,2-d]pyrimidin-4-yl]morpholine(also known as GDC 0941 and described in PCT Publication Nos. WO09/036,082 and WO 09/055,730),2-Methyl-2-[4-[3-methyl-2-oxo-8-(quinolin-3 -yl)-2,3 -dihydroimidazo[4,5-c]quinolin-1-yl]phenyl]propionitrile (also known as BEZ 235 orNVP-BEZ 235, and described in PCT Publication No. WO 06/122806),(S)-1-(4-((2-(2-aminopyrimidin-5-yl)-7-methyl-4-morpholinothieno[3,2-d]pyrimidin-6-yl)methyl)piperazin-1-yl)-2-hydroxypropan-1-one(described in PCT Publication No. WO 2008/070740), LY294002(2-(4-Morpholinyl)-8-phenyl-4H-1-benzopyran-4-one available from AxonMedchem), PI 103 hydrochloride(3-[4-(4-morpholinylpyrido-[3′,2′:4,5]furo[3,2-d]pyrimidin-2-yl]phenolhydrochloride available from Axon Medchem), PIK 75(N′-[(1E)-(6-bromoimidazo[1,2-a]pyridin-3-yl)methylene]-N,2-dimethyl-5-nitrobenzenesulfono-hydrazide hydrochloride available from AxonMedchem), PIK 90(N-(7,8-dimethoxy-2,3-dihydro-imidazo[1,2-c]quinazolin-5-yl)-nicotinamideavailable from Axon Medchem), GDC-0941 bismesylate(2-(1H-Indazol-4-yl)-6-(4-methanesulfonyl-piperazin-1-ylmethyl)-4-morpholin-4-yl-thieno[3,2-d]pyrimidinebismesylate available from Axon Medchem), AS-252424 (5-[1-[5-(4-Fluoro-2-hydroxy-phenyl)-furan-2-yl]-meth-(Z)-ylidene]-thiazolidine-2,4-dioneavailable from Axon Medchem), and TGX-221(7-Methyl-2-(4-morpholinyl)-9-[1-(phenylamino)ethyl]-4H-pyrido-[1,2-a]pyrimidin-4-oneavailable from Axon Medchem), XL-765, and XL-147. Other PI3K inhibitorsinclude demethoxyviridin, perifosine, CAL101, PX-866, BEZ235, SF1126,INK1117, IPI-145, BKM120, XL147, XL765, Palomid 529, GSK1059615,ZSTK474, PWT33597, IC87114, TG100-115, CAL263, PI-103, GNE-477,CUDC-907, and AEZS-136.

AKT inhibitors include, but are not limited to, Akt-1-1 (inhibits Aktl)(Barnett et al. (2005) Biochem. 1, 385 (Pt. 2), 399-408); Akt-1-1,2(inhibits AK1 and 2) (Barnett et al. (2005) Biochem. J. 385 (Pt. 2),399-408); API-59CJ-Ome (e.g., Jin et al. (2004) Br J. Cancer 91,1808-12); 1-H-imidazo[4,5-c]pyridinyl compounds (e.g., WO05011700);indole-3-carbinol and derivatives thereof (e.g., U.S. Pat. No.6,656,963; Sarkar and Li (2004) Jl Nutr. 134(12 Suppl), 3493S-3498S);perifosine (e.g., interferes with Akt membrane localization;Dasmahapatra et al. (2004) Clin. Cancer Res. 10(15), 5242-52, 2004);phosphatidylinositol ether lipid analogues (e.g., Gills and Dennis(2004) Expert. Opin. Investig. Drugs 13, 787-97); and triciribine (TCNor API-2 or NCI identifier: NSC 154020; Yang et al. (2004) Cancer Res.64, 4394-9).

TOR inhibitors include, but are not limited to, AP-23573, CCI-779,everolimus, RAD-001, rapamycin, temsirolimus, ATP-competitiveTORC1/TORC2 inhibitors, including PI-103, PP242, PP30 and Torin 1. OtherTOR inhibitors in FKBP12 enhancer; rapamycins and derivatives thereof,including: CCI-779 (temsirolimus), RAD001 (Everolimus; WO 9409010) andAP23573; rapalogs, e.g. as disclosed in WO 98/02441 and WO 01/14387,e.g. AP23573, AP23464, or AP23841; 40-(2-hydroxyethyl)rapamycin,40-[3-hydroxy(hydroxymethyl)methylpropanoate]-rapamycin (also calledCC1779), 40-epi-(tetrazolyt)-rapamycin (also called ABT578),32-deoxorapamycin, 16-pentynyloxy-32(S)-dihydrorapanycin, and otherderivatives disclosed in WO 05005434; derivatives disclosed in U.S. Pat.No. 5,258,389, WO 94/090101, WO 92/05179, U.S. Pat. Nos. 5,118,677,5,118,678, 5,100,883, 5,151,413, 5,120,842, WO 93/111130, WO 94/02136,WO 94/02485, WO 95/14023, WO 94/02136, WO 95/16691, WO 96/41807, WO96/41807 and U.S. Pat. No. 5,256,790; phosphorus-containing rapamycinderivatives (e.g., WO 05016252); 4H-1-benzopyran-4-one derivatives(e.g., U.S. Provisional Application No. 60/528,340).

MC₁-1 inhibitors include, but are not limited to, AMG-176, MIK665, and563845. The myeloid cell leukemia-1 (MCL-1) protein is one of the keyanti-apoptotic members of the B-cell lymphoma-2 (BCL-2) protein family.Over-expression of MCL-1 has been closely related to tumor progressionas well as to resistance, not only to traditional chemotherapies butalso to targeted therapeutics including BCL-2 inhibitors such asABT-263.

KRAS^(G12C) inhibitors can also be used in combination with SHP2inhibitors in the present invention. SHP2 inhibitors that can be used inthe present combinations include, but are not limited to, SHP099, andRMC-4550 or RMC-4630, from Revolutions Medicines in Redwood City, Calif.

Proteasome inhibitors include, but are not limited to,Kyprolis®(carfilzomib), Velcade®(bortezomib), and oprozomib.

Immune therapies include, but are not limited to, anti-PD-1 agents,anti-PDL-1 agents, anti-CTLA-4 agents, anti-LAG1 agents, and anti-OX40agents.

Monoclonal antibodies include, but are not limited to, Darzalex®(daratumumab), Herceptin® (trastuzumab), Avastin® (bevacizumab),Rittman® (rituximab), Lucentis® (ranibizumab), and Eylea® (aflibercept).

Immunomodulatory agents (IMiDs) are a class of immunomodulatory drugs(drugs that adjust immune responses) containing an imide group. The IMiDclass includes thalidomide and its analogues (lenalidomide,pomalidomide, and apremilast).

Anti-PD-1 inhibitors, including but not limited to antibodies include,but are not limited to, pembrolizumab (Keytruda®) and nivolumab(Opdive). Exemplary anti-PD-1 antibodies and methods for their use aredescribed by Goldberg et al., Blood 110(1):186-192 (2007), Thompson etal., Clin. Cancer Res. 13(6):1757-1761 (2007), and Korman et al.,International Application No. PCT/JP2006/309606 (publication no. WO2006/121168 A1), each of which are expressly incorporated by referenceherein. include: Yervoy™ (ipilimumab) or Tremelimumab (to CTLA-4),galiximab (to B7.1), BMS-936558 (to PD-1), MK-3475 (to PD-1), AMP224 (toB7DC), BMS-936559 (to B7-H1), MPDL3280A (to B7-H1), MEDI-570 (to ICOS),AMG557 (to B7H2), MGA271 (to B7H3), IMP321 (to LAG-3), BMS-663513 (toCD137), PF-05082566 (to CD137), CDX-1127 (to CD27), anti-OX40(Providence Health Services), huMAbOX40L (to OX40L), Atacicept (toTACI), CP-870893 (to CD40), Lucatumumab (to CD40), Dacetuzumab (toCD40), Muromonab-CD3 (to CD3), Ipilumumab (to CTLA-4). Immune therapiesalso include genetically engineered T-cells (e.g., CAR-T cells) andbispecific antibodies (e.g., BiTEs).

GITR agonists include, but are not limited to, GITR fusion proteins andanti-GITR antibodies (e.g., bivalent anti-GITR antibodies), such as, aGITR fusion protein described in U.S. Pat. No. 6,111,090box.c, EuropeanPatent No.: 090505B1, U.S. Pat. No. 8,586,023, PCT Publication Nos.: WO2010/003118 and 2011/090754, or an anti-GITR antibody described, e.g.,in U.S. Pat. No. 7,025,962, European Patent No.: 1947183B1, U.S. Pat.Nos. 7,812,135, 8,388,967, 8,591,886, European Patent No.: EP 1866339,PCT Publication No.: WO 2011/028683, PCT Publication No.: WO2013/039954, PCT Publication No.: WO2005/007190, PCT Publication No.: WO2007/133822, PCT Publication No.: WO2005/055808, PCT Publication No.: WO99/40196, PCT Publication No.: WO 2001/03720, PCT Publication No.:WO99/20758, PCT Publication No.: WO2006/083289, PCT Publication No.: WO2005/115451, U.S. Pat. No. 7,618,632, and PCT Publication No.: WO2011/051726.

The compounds described herein can be used in combination with theagents disclosed herein or other suitable agents, depending on thecondition being treated. Hence, in some embodiments the one or morecompounds of the disclosure will be co-administered with other agents asdescribed above. When used in combination therapy, the compoundsdescribed herein are administered with the second agent simultaneouslyor separately. This administration in combination can includesimultaneous administration of the two agents in the same dosage form,simultaneous administration in separate dosage forms, and separateadministration. That is, a compound described herein and any of theagents described above can be formulated together in the same dosageform and administered simultaneously. Alternatively, a compound of thedisclosure and any of the agents described above can be simultaneouslyadministered, wherein both the agents are present in separateformulations. In another alternative, a compound of the presentdisclosure can be administered just followed by and any of the agentsdescribed above, or vice versa. In some embodiments of the separateadministration protocol, a compound of the disclosure and any of theagents described above are administered a few minutes apart, or a fewhours apart, or a few days apart.

As one aspect of the present invention contemplates the treatment of thedisease/conditions with a combination of pharmaceutically activecompounds that may be administered separately, the invention furtherrelates to combining separate pharmaceutical compositions in kit form.The kit comprises two separate pharmaceutical compositions: a compoundof the present invention, and a second pharmaceutical compound. The kitcomprises a container for containing the separate compositions such as adivided bottle or a divided foil packet. Additional examples ofcontainers include syringes, boxes, and bags. In some embodiments, thekit comprises directions for the use of the separate components. The kitform is particularly advantageous when the separate components arepreferably administered in different dosage forms (e.g., oral andparenteral), are administered at different dosage intervals, or whentitration of the individual components of the combination is desired bythe prescribing health care professional.

All patents and other publications recited herein are herebyincorporated by reference.

The processes presented below illustrate specific embodiments of thepresent invention. These processes are meant to be representative andare not intended to limit the scope of the claims in any manner.

Representative Examples of the Invention

The following intermediate compounds of6-Fluoro-7-(2-fluoro-6-hydroxyphenyl)-1-(4-methyl-2-(2-propanyl)-3-pyridinyl)-4-((2S)-2-methyl-4-(2-propenoyl)-1-piperazinyl)pyrido[2,3-d]pyrimidin-2(1H)-oneare representative examples of the invention and are not intended to beconstrued as limiting the scope of the present invention.

A synthesis of Compound 9 and the relevant intermediates is described inU.S. Ser. No. 15/984,855, filed May 21, 2018, which claims priority toand the benefit claims the benefit of U.S. Provisional Application No.62/509,629, filed on May 22, 2017, which are incorporated herein byreference in their entireties for all purposes.6-Fluoro-7-(2-fluoro-6-hydroxyphenyl)-1-(4-methyl-2-(2-propanyl)-3-pyridinyl)-4-((2S)-2-methyl-4-(2-propenoyl)-1-piperazinyl)pyrido[2,3-d]pyrimidin-2(1H)-onewas prepared using the following process, in which the isomers of thefinal product were isolated via chiral chromatography.

Step 1: 2,6-Dichloro-5-fluoronicotinamide (Intermediate S). To a mixtureof 2,6-dichloro-5-fluoro-nicotinic acid (4.0 g, 19.1 mmol, AstaTechInc., Bristol, Pa.) in dichloromethane (48 mL) was added oxalyl chloride(2M solution in DCM, 11.9 mL, 23.8 mmol), followed by a catalytic amountof DMF (0.05 mL). The reaction was stirred at room temperature overnightand then was concentrated. The residue was dissolved in 1,4-dioxane (48mL) and cooled to 0° C. Ammonium hydroxide solution (28.0-30% NH3 basis,3.6 mL, 28.6 mmol) was added slowly via syringe. The resulting mixturewas stirred at 0° C. for 30 min and then was concentrated. The residuewas diluted with a 1:1 mixture of EtOAc/Heptane and agitated for 5 min,then was filtered. The filtered solids were discarded, and the remainingmother liquor was partially concentrated to half volume and filtered.The filtered solids were washed with heptane and dried in areduced-pressure oven (45° C.) overnight to provide2,6-dichloro-5-fluoronicotinamide. ¹H NMR (400 MHz, DMSO-d₆) δ ppm 8.23(d, J=7.9 Hz, 1H) 8.09 (br s, 1H) 7.93 (br s, 1H). m/z (ESI, +ve ion):210.9 (M+H)⁺.

Step 2:2,6-Dichloro-5-fluoro-N-((2-isopropyl-4-methylpyridin-3-yl)carbamoyl)nicotinamide.To an ice-cooled slurry of 2,6-dichloro-5-fluoronicotinamide(Intermediate S, 5.0 g, 23.9 mmol) in THF (20 mL) was added oxalylchloride (2 M solution in DCM, 14.4 mL, 28.8 mmol) slowly via syringe.The resulting mixture was heated at 75° C. for 1 h, then heating wasstopped, and the reaction was concentrated to half volume. After coolingto 0° C., THF (20 mL) was added, followed by a solution of2-isopropyl-4-methylpyridin-3-amine (Intermediate R, 3.59 g, 23.92 mmol)in THF (10 mL), dropwise via cannula. The resulting mixture was stirredat 0° C. for 1 h and then was quenched with a 1:1 mixture of brine andsaturated aqueous ammonium chloride. The mixture was extracted withEtOAc (3×) and the combined organic layers were dried over anhydroussodium sulfate and concentrated to provide2,6-dichloro-5-fluoro-N-((2-isopropyl-4-methylpyridin-3-yl)carbamoyl)nicotinamide.This material was used without further purification in the followingstep. m/z (ESI, +ve ion): 385.1(M+H)⁺.

Step 3:7-Chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)pyrido[2,3-d]pyrimidine-2,4(1H,3H)-dione.To an ice-cooled solution of2,6-dichloro-5-fluoro-N-((2-isopropyl-4-methylpyridin-3-yl)carbamoyl)nicotinamide(9.2 g, 24.0 mmol) in THF (40 mL) was added KHMDS (1 M solution in THF,50.2 mL, 50.2 mmol) slowly via syringe. The ice bath was removed and theresulting mixture was stirred for 40 min at room temperature. Thereaction was quenched with saturated aqueous ammonium chloride andextracted with EtOAc (3×). The combined organic layers were dried overanhydrous sodium sulfate and concentrated. The residue was purified bysilica gel chromatography (eluent: 0-50% 3:1 EtOAc-EtOH/heptane) toprovide7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)pyrido[2,3-d]pyrimidine-2,4(1H,3H)-dione.¹H NMR (400 MHz, DMSO-d₆) δ ppm 12.27 (br s, 1H), 8.48-8.55 (m, 2H),7.29 (d, J=4.8 Hz, 1H), 2.87 (quin, J=6.6 Hz, 1H), 1.99-2.06 (m, 3H),1.09 (d, J=6.6 Hz, 3H), 1.01 (d, J=6.6 Hz, 3H). ¹⁹F NMR (376 MHz,DMSO-d₆) δ: −126.90 (s, 1F). m/z (ESI, +ve ion): 349.1 (M+H)⁺.

Step 4:4,7-Dichloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)pyrido[2,3-d]pyrimidin-2(1H)-one.To a solution of7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)pyrido[2,3-d]pyrimidine-2,4(1H,3H)-dione(4.7 g, 13.5 mmol) and DIPEA (3.5 mL, 20.2 mmol) in acetonitrile (20 mL)was added phosphorus oxychloride (1.63 mL, 17.5 mmol), dropwise viasyringe. The resulting mixture was heated at 80° C. for 1 h, and thenwas cooled to room temperature and concentrated to provide4,7-dichloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)pyrido[2,3-d]pyrimidin-2(1H)-one. This material was used without furtherpurification in the following step. m/z (ESI, +ve ion): 367.1 (M+H)⁺.

Step 5: (S)-tert-Butyl4-(7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-2-oxo-1,2-dihydropyrido[2,3-d]pyrimidin-4-yl)-3-methylpiperazine-1-carboxylate.To an ice-cooled solution of4,7-dichloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)pyrido[2,3-d]pyrimidin-2(1H)-one(13.5 mmol) in acetonitrile (20 mL) was added DIPEA (7.1 mL, 40.3 mmol),followed by (S)-4-N-Boc-2-methyl piperazine (3.23 g, 16.1 mmol,Combi-Blocks, Inc., San Diego, Calif., USA). The resulting mixture waswarmed to room temperature and stirred for 1 h, then was diluted withcold saturated aqueous sodium bicarbonate solution (200 mL) and EtOAc(300 mL). The mixture was stirred for an additional 5 min, the layerswere separated, and the aqueous layer was extracted with more EtOAc(1×). The combined organic layers were dried over anhydrous sodiumsulfate and concentrated. The residue was purified by silica gelchromatography (eluent: 0-50% EtOAc/heptane) to provide (S)-tent-butyl4-(7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-2-oxo-1,2-dihydropyrido[2,3-d]pyrimidin-4-yl)-3-methylpiperazine-1-carboxylate.m/z (ESI, +ve ion): 531.2 (M+H)⁺.

Step 6: (3S)-tert-Butyl4-(6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-1-(2-isopropyl-4-methylpyridin-3-yl)-2-oxo-1,2-dihydropyrido[2,3-d]pyrimidin-4-yl)-3-methylpiperazine-1-carboxylate.A mixture of (S)-tent-butyl4-(7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)-2-oxo-1,2-dihydropyrido[2,3-d]pyrimidin-4-yl)-3-methylpiperazine-1-carboxylate(4.3 g, 8.1 mmol), potassium trifluoro(2-fluoro-6-hydroxyphenyl)borate(Intermediate Q, 2.9 g, 10.5 mmol), potassium acetate (3.2 g, 32.4 mmol)and [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II), complexwith dichloromethane (661 mg, 0.81 mmol) in 1,4-dioxane (80 mL) wasdegassed with nitrogen for 1 min. De-oxygenated water (14 mL) was added,and the resulting mixture was heated at 90° C. for 1 h. The reaction wasallowed to cool to room temperature, quenched with half-saturatedaqueous sodium bicarbonate, and extracted with EtOAc (2×) and DCM (1×).The combined organic layers were dried over anhydrous sodium sulfate andconcentrated. The residue was purified by silica gel chromatography(eluent: 0-60% 3:1 EtOAc-EtOH/heptane) to provide (3S)-tert-butyl4-(6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-1-(2-isopropyl-4-methylpyridin-3-yl)-2-oxo-1,2-dihydropyrido[2,3-d]pyrimidin-4-yl)-3-methylpiperazine-1-carboxylate.¹H NMR (400 MHz, DMSO-d₆) δ ppm 10.19 (br s, 1H), 8.38 (d, J=5.0 Hz,1H), 8.26 (dd, J=12.5, 9.2 Hz, 1H), 7.23-7.28 (m, 1H), 7.18 (d, J=5.0Hz, 1H), 6.72 (d, J=8.0 Hz, 1H), 6.68 (t, J=8.9 Hz, 1 H), 4.77-4.98 (m,1H), 4.24 (br t, J=14.2 Hz, 1H), 3.93-4.08 (m, 1H), 3.84 (br d, J=12.9Hz, 1H), 3.52-3.75 (m, 1H), 3.07-3.28 (m, 1H), 2.62-2.74 (m, 1H),1.86-1.93 (m, 3H), 1.43-1.48 (m, 9H), 1.35 (dd, J=10.8, 6.8 Hz, 3H),1.26-1.32 (m, 1H), 1.07 (dd, J=6.6, 1.7 Hz, 3H), 0.93 (dd, J=6.6, 2.1Hz, 3H). ¹⁹F NMR (376 MHz, DMSO-d₆)δ: −115.65 (s, 1F), −128.62 (s, 1F).m/z (ESI, +ve ion): 607.3 (M+H)⁺.

Step 7:6-Fluoro-7-(2-fluoro-6-hydroxyphenyl)-1-(4-methyl-2-(2-propanyl)-3-pyridinyl)-4-(2S)-2-methyl-4-(2-propenoyl)-1-piperazinyl)pyrido[2,3-d]pyrimidin-2(1H)-one.Trifluoroacetic acid (25 mL, 324 mmol) was added to a solution of(3S)-tert-butyl4-(6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-1-(2-isopropyl-4-methylpyridin-3-yl)-2-oxo-1,2-dihydropyrido[2,3-d]pyrimidin-4-yl)-3-methylpiperazine-1-carboxylate(6.3 g, 10.4 mmol) in DCM (30 mL). The resulting mixture was stirred atroom temperature for 1 h and then was concentrated. The residue wasdissolved in DCM (30 mL), cooled to 0° C., and sequentially treated withDIPEA (7.3 mL, 41.7 mmol) and a solution of acryloyl chloride (0.849 mL,10.4 mmol) in DCM (3 mL; added dropwise via syringe). The reaction wasstirred at 0° C. for 10 min, then was quenched with half-saturatedaqueous sodium bicarbonate and extracted with DCM (2×). The combinedorganic layers were dried over anhydrous sodium sulfate andconcentrated. The residue was purified by silica gel chromatography(eluent: 0-100% 3:1 EtOAc-EtOH/heptane) to provide6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-1-(4-methyl-2-(2-propanyl)-3-pyridinyl)-4-(2S)-2-methyl-4-(2-propenoyl)-1-piperazinyl)pyrido[2,3-d]pyrimidin-2(1H)-one.¹H NMR (400 MHz, DMSO-d₆) δ ppm 10.20 (s, 1H), 8.39 (d, J=4.8 Hz, 1H),8.24-8.34 (m, 1H), 7.23-7.32 (m, 1H), 7.19 (d, J=5.0 Hz, 1H), 6.87 (td,J=16.3, 11.0 Hz, 1H), 6.74 (d, J=8.6 Hz, 1H), 6.69 (t, J=8.6 Hz, 1H),6.21 (br d, J=16.2 Hz, 1H), 5.74-5.80 (m, 1H), 4.91 (br s, 1H),4.23-4.45 (m, 2H), 3.97-4.21 (m, 1H), 3.44-3.79 (m, 2H), 3.11-3.31 (m,1H), 2.67-2.77 (m, 1H), 1.91 (s, 3H), 1.35 (d, J=6.8 Hz, 3H), 1.08 (d,J=6.6 Hz, 3H), 0.94 (d, J=6.8 Hz, 3H). ¹⁹F NMR (376 MHz, DMSO-d₆) δ ppm-115.64 (s, 1F), −128.63 (s, 1F). m/z (ESI, +ve ion): 561.2 (M+H)⁺.

The present invention comprises the following steps wherein theresolution of the rac-Dione in Steps 4 and 5 promotes the successfulseparation of the atropisomers:

Process Description Step 1

MW Equivalents/ Material CAS # (g/mol) Volumes Moles Theoretical2,6-dichloro-5-fluoro-3- 82671-06-5 209.99   1.0 equiv. 119.1 25 kgpyridinecarboxylic acid DCM 74-09-2 84.93 16.51 equiv. 2354.9 200 kg DMF68-12-2 73.09 0.068 equiv. 8.1 592 g (627 mL) Oxalyl Chloride 79-37-8126.93  1.25 equiv. 148.9 18.9 kg Ammonium Hydroxide 1336-21-6 35.05   5 equiv. 595.5 40.2 L Water 7732-18-5 18.02 N/A N/A 261 L

To a solution of 2,6-dichloro-5-fluoro-3-pyridinecarboxylic acid(Compound 1) (25 kg; 119.1mol) in dichloromethane (167 kg) and DMF (592g) was added Oxalyl chloride (18.9 kg; 148.9 mol) while maintaining aninternal temp between 15-20° C. Additional dichloromethane (33 kg) wasadded as a rinse and the reaction mixture stirred for 2 h. The reactionmixture is cooled then quenched with ammonium hydroxide (40.2 L; 595.5mol) while maintaining internal temperature 0±10° C. The resultingslurry was stirred for 90 min then the product collected by filtration.The filtered solids were washed with DI water (3×87 L) and dried toprovide 2,6-dichloro-5-fluoronicotinamide (Compound 2).

Step 2

MW Equivalents Material CAS# (g/mol) Volumes Moles Theoretical Amide113237-20-0 209.99 1.0 equiv. 77.8 16.27 kg (2,6-dichloro-5-fluoronicotinamide) Oxalyl Chloride 79-37-8 126.93 1.2 equiv. 93.8 11.9kg (7.9 L) 730.7 kg Dichloromethane 75-09-2 84.93 N/A N/A (551.5 L)Aniline DCM Solution 1698293-93-4 150.22 1.1 equiv. 85.9 12.9 kg(Aniline 2-isopropyl-4- contained wt) methylpyridin-3- amine

In reactor A, a solution of 2,6-dichloro-5-fluoronicotinamide (Compound2) (16.27 kg; 77.8 mol) in dichloromethane (359.5 kg) was added oxalylchloride (11.9 kg; 93.8 mol) while maintaining temp <25° C. for 75 min.The resulting solution was then headed to 40° C.±3° C. and aged for 3 h.Using vacuum, the solution was distilled to remove dichloromethane untilthe solution was below the agitator. Dichloromethane (300 kg) was thenadded and the mixture cooled to 0±5° C. To a clean, dry reactor (reactorB) was added,2-isopropyl-4-methylpyridin-3-amine (ANILINE) (12.9 kg;85.9 mol) followed by dichloromethane (102.6 kg). The ANILINE solutionwas azeodried via vacuum distillation while maintaining a internaltemperature between 20-25°), replacing with additional dichloromethaneuntil the solution was dry by KF analysis (limit <0.05%). The solutionvolume was adjusted to approx. 23 L volume with dichloromethane. Thedried ANILINE solution was then added to reactor A while maintaining aninternal temperature of 0±5° C. throughout the addition. The mixture wasthen heated to 23° C. and aged for 1 h. the solution was polish filteredinto a clean reactor to afford2,6-dichloro-5-fluoro-N-((2-isopropyl-4-methylpyridin-3-yl)carbamoyl)nicotinamide(Compound 3) as a solution in DCM and used directly in the next step.

Step 3

MW Equivalents/ Material CAS # (g/mol) Volumes Moles Theoretical Urea,solution in DCM N/A 385.22 1.0 equiv. 38.9 208.3 kg2,6-dichloro-5-fluoro- (15 kg N-{[4-methyl-2- contained weight)(propan-2-yl)pyridin-3- yl]carbamoyl}pyridine- 3-carboxamide2-methyltetrahydrofuran 96-47-9 86.13 N/A N/A 308 kg (358 L) Sodiumtert-butoxide 865-48-5 96.11 2.0 equiv 97.8 9.4 kg Ammonium Chloride12125-02-9 53.49 N/A 430 23.0 kg Hydrochloric Acid 7467-01-0 36.46 N/A41 1.6 kg Magnesium Sulfate 7487-88-9 120.37 N/A 195 23.5 kg SodiumChloride 7647-14-5 58.44 N/A 282 16.5 kg Heptane 142-82-5 100.21 N/A N/A94 L 10% citric acid 75 kg

A dichloromethane solution of2,6-dichloro-5-fluoro-N-{[4-methyl-2-(propan-2-yl)pyridin-3-yl]carbamoyl}pyridine-3-carboxamide(UREA(Compound 3)) (15 kg contained; 38.9 mol) was solvent exchangedinto 2-MeTHF using vacuum distillation while maintaining internaltemperature of 20-25° C. The reactor volume was adjusted to 40 L andthen additional 2-MeTHF was charged (105.4 kg). Sodium t-butoxide wasadded (9.4 kg; 97.8 mol) while maintaining 5-10° C. The contents wherewarmed to 23° C. and stirred for 3 h. The contents where then cooled to0-5C and ammonium chloride added (23.0 kg; 430 mol) as a solution in 60L of DI water. The mixture was warmed to 20 C and DI water added (15 L)and further aged for 30 min. Agitation was stopped and the layersseparated. The aqueous layer was removed and to the organic layer wasadded DI water(81.7 L). A mixture of conc HCl (1.5 kg) and water (9 L)was prepared then added to the reactor slowly until pH measured between4-5. The layers were separated, and the aqueous layer back extractedusing 2-MeTHF (42.2 kg). The two organic layers combined and washed witha 10% citric acid solution (75 kg) followed by a mixture of water (81.7L) and saturated NaCl (19.8 kg). The organic layer was then washed withsaturated sodium bicarbonate (75 kg) repeateding if necessary to achievea target pH of ≥7.0 of the aqueous. The organic layer was washed againwith brine (54.7 kg) and then dried over magnesium sulfate (5 kg). Themixture was filtered to remove magnesium sulfate rinsing the filteredbed with 2-MeTHF (49.2 kg). The combined filtrate and washes wheredistilled using vacuum to 40 L volume. The concentrated solution washeated to 55° C. and heptane (10-12 kg) slowly added until cloud point.The solution was cooled to 23° C. over 2 h then heptane (27.3 kg) wasadded over 2 h. The product slurry was aged for 3 h at 20-25° C. thenfiltered and washed with a mixture of 2-MeTHF (2.8 kg) and heptane (9kg). The product was dried using nitrogen and vacuum to afford solid7-chloro-6-fluoro-1-(2-isopropyl-4-methylpyridin-3-yl)pyrido[2,3-d]pyrimidine-2,4(1H,3H)-dione(rac-DIONE (Compound 4)).

Step 4

MW Equivalents/ Material CAS # (g/mol) Volumes Moles TheoreticalRac-dione N/A 348.76 1.0 (+)-2,3-dibenzoyl-D- 17026-42-5 358.30 2.0tartaric acid 2-methyltetrahydrofuran 96-47-9  86.13 7.0 heptane142-82-5 100.21 2.0 heptane 142-82-5 100.21 3.0 2-methyltetrahydrofuran96-47-9  86.13 4.0 heptane 142-82-5 100.21 2.0

To a vessel, an agitated suspension of Compound 4, (1.0 eq.) in2-methylterahydrofuran (7.0 L/kg) was added (+)-2,3-dibenzoyl-D-tartaricacid (2.0 eq.) under an atmosphere of nitrogen. 2-MeTHF is chiral, butit is used as a racemic mixture. The different enantiomers of 2-MeTHFare incorporated randomly into the co-crystal. The resulting suspensionwas warmed to 75° C. and aged at 75° C. until full dissolution wasobserved (≤30 mins.). The resulting solution was polish filtered at 75°C. into a secondary vessel. To the polish filtered solution was chargedn-Heptane (2.0 L/kg) at a rate that maintained the internal temperatureabove 65° C. The solution was then cooled to 60° C., seeded withcrystals (0.01 kg/kg) and allowed to age for 30 minutes. The resultingsuspension was cooled to 20° C. over 4 hours and then sampled for chiralpurity analysis by HPLC. To the suspension, n-Heptane (3.0 L/kg) wascharged and then aged for 4 hours at 20° C. under an atmosphere ofnitrogen. The suspension was filtered, and the isolated solids werewashed two times with (2:1) n-Heptane:2-methyltetrahydrofuran (3.0L/kg). The material was dried with nitrogen and vacuum to affordM-Dione:DBTA: Me-THF complex (Compound 4a).

Step 5

MW Equivalents/ Material CAS # (g/mol) Volumes Moles TheoreticalM-Dione/DBTA/Me- N/A 1228.08  1.0 74.2 46.9 kg THF cocrystal (25.9 kgcorrected for M- dione) Methyl tert-butyl ether 1634-04-4  88.15 45.017593 2100 L Disodium hydrogen 7558-79-4 141.96  2.0 148.4 21.1 kgphosphate USP purified water As needed Magnesium sulfate 7487-88-9120.37 N/A N/A 25 kg Heptane 142-82-5 100.20 60.0 19322 2835 L

To vessel A, a suspension of disodium hydrogen phosphate (21.1 kg, 2.0equiv) in DI water (296.8 L, 6.3 L/kg) was agitated until dissolutionwas observed (>30 min.). To vessel B, a suspension of the M-Dione:DBTA:Me-THF complex (Composition 4a)[46.9 kg (25.9 kg corrected for M-dione,1.0 equiv.)] in methyl tert-butyl ether (517.8 L, 11.0 L/kg) wasagitated for 15 to 30 minutes. The resulting solution from vessel A wasadded to vessel B, and then the mixture was agitated for more than 3hours. The agitation was stopped, and the biphasic mixture was left toseparate for more than 30 minutes. The lower aqueous phase was removedand then back extracted with methyl tert-butyl ether (77.7 L, 1.7 L/kg).The organic phases were combined in vessel B and dried with magnesiumsulfate (24.8 kg, 0.529 kg/kg). The resulting suspension from vessel Bwas agitated for more than three hours and then filtered into vessel C.To vessel B, a methyl tert-butyl ether (46.9 L, 1.0 L/kg) rinse wascharged and then filtered into vessel C. The contents of vessel C werecooled to 10° C. and then distilled under vacuum while slowly beingwarmed to 35° C. Distillation was continued until 320-350 kg (6.8-7.5kg/kg) of methyl tert-butyl ether was collected. After cooling thecontents of vessel C to 20° C., n-Heptane (278.7 L, 5.9 L/kg) wascharged over one hour and then distilled under vacuum while slowly beingwarmed to 35° C. Distillation was continued until a 190-200 kg (4.1-4.3kg/kg) mixture of methyl tert-butyl ether and n-Heptane was collected.After cooling the contents of vessel C to 20° C., n-Heptane (278.7 L,5.9 L/kg) was charged a second time over one hour and then distilledunder vacuum while slowly being warmed to 35° C. Distillation wascontinued until a 190-200 kg (4.1-4.3 kg/kg) mixture of methyltert-butyl ether and n-Heptane was collected. After cooling the contentsof vessel C to 20° C., n-Heptane (195.9 L, 4.2 L/kg) was charged a thirdtime over one hour and then sampled for solvent composition by GCanalysis. The vessel C suspension continued to agitate for more than onehour. The suspension was filtered, and then washed with a n-Heptane(68.6 L, 1.5 L/kg) rinse from vessel C. The isolated solids were driedat 50° C., and a sample was submitted for stock suitability. Afforded7-chloro-6-fluoro-(1M)-1-[4-methyl-2-(propan-2-yl)pyridin-3-yl]pyrido[2,3-d]pyrimidine-2,4(1H,3H)-dione(M-DIONE) Compound 5M.

The first-generation process highlighted above has been successfullyscaled on 200+ kg of rac-dione starting material (Compound 5). In thisprocess, seeding the crystallization with the thermodynamically-stablerac-dione crystal form (which exhibits low solubility) would cause abatch failure. Based on our subsequent studies, we found that increasingthe DBTA equivalents and lowering the seed temperature by adjustingheptane charge schedule improves robustness of the process. The improvedprocess is resistant to the presence of the thermodynamically-stablerac-dione crystal form and promotes successful separation ofatropisomers. Subsequent batches will incorporate the improved processfor large scale manufacture.

Step 6

MW Equivalents/ Material CAS # (g/mol) Volumes Moles Theoretical M-DIONEN/A 348.76   1 equiv. 9.8 3.7 kg Toluene 108-88-3 92.14 N/A 375 34.6 kg(40 L) Phosphoryl chloride 10025-87-3 153.33 1.2 equiv. 11.7 1.8 kg (1.1L) N,N- 7087-68-5 129.24 3.0 equiv. 29.4 3.8 kg (5.1 L)Diisopropylethylamine (s)-1-Boc-3- 147081-29-6 200.28 1.1 equiv. 10.82.214 kg methylpiperazine Sodium bicarbonate 144-55-8 84.01 N/A N/A 973g Dichloromethane 75-09-2 84.93 N/A 871 74 kg (55.6 L) Sodium Chloride7647-14-5 58.44 N/A 103 6.0 kg Ethyl acetate 141-78-6 88.11 N/A 288 25.4kg (28.2 L)

7-chloro-6-fluoro-(1M)-1-[4-methyl-2-(propan-2-yl)pyridin-3-yl]pyrido[2,3-d]pyrimidine-2,4(1H,3H)-dione(M-DIONE) (3.7 kg; 9.8 mol) was combined in reactor (A) with 10.5 kg oftoluene and distilled down to an oil to remove water while maintaining aset point of 45° C. Toluene (21 kg) was added to the residue and themixture stirred for 30 min at 40-45° C. The contents where cooled to 22°C. then phosphoryl chloride (1.8 kg; 11.7 mol) added. The mixture wascooled to 0-5° C. before adding N,N-Diisopropylethylamine (2.5 kg; 19.34mol) while maintaining a temperature <5° C. The solution was aged for 3h at 22° C. In a separate reactor (B), (s)-1-boc-3-methylpiperazine(2.21 kg; 10.8 mol) and N,N-diisopropylethylamine (1.26 kg; 9.75 mol))where combined in toluene (6 kg) and then charged to reactor (A) whilemaintaining <25° C. The reaction mixture was aged for 15 min at 22 Cthen quenched with sodium bicarbonate (973 g) in water (12.9 L) whilemaintaining a temperature <25 C. The mixture was stirred for 30 min thenDCM (36.8 kg) added while continuing to stir for 1 h. The layers wereallowed to separate, and the lower organic layer drained to reactor (C).The aqueous layer in reactor (A) was back extracted using DCM (18.4 kg)and the combined organic layers washed with brine solution (6.0 kg NaCl;16.5 kg DI water). The organic layer was distilled under atmosphericpressure maintaining an internal temperature between 45-55 C. DCM isreplaced during the distillation to azeotropically dry the solution.Following the distillation, the solution volume was adjusted to 19 Lusing DCM. The solution was cooled to 30 C and polish filtered. Thefiltrate was combined with ethyl acetate (8.5 kg) and then distilled atatmospheric pressure until 11-13 kg is collected in the receiver. Thesolution was seeded with 30 g of authentic product and aged for 1 h at25-30° C. then further distilled under atmospheric pressure at 45-55 Cinternal temperature until 8.2 kg of distillate had been collected. Theslurry was cooled to 22° C. and aged overnight then further cooled to0-5° C. The product was collected by filtration and washed twice usingethyl acetate (4.2 kg each). The cake was dried with nitrogen and vacuumto afford tent-butyl(3S)-4-{7-chloro-6-fluoro-(1M)-1-[4-methyl-2-(propan-2-yl)pyridin-3-yl]-2-oxo-1,2-dihydropyrido[2,3-d]pyrimidin-4-yl}-3-methylpiperazine-1-carboxylate(Compound 6, PIPAZOLINE).

Step 7

MW Equivalents/ Material CAS # (g/mol) Volumes Moles Theoretical [1,1′-72287- 731.714  0.020 1.01 0.74 kg Bis(diphenylphosphino) 26-4ferrocene] dichloropalladium(II) Dichloromethane 75-09-2 84.93 N/A 400kg 1,4-Dioxane 123-91-1 88.1052 5.0 N/A 168 kgEthylenediaminetetraacetic 6381-92-6 336.207 1.0 45.2 15.2 kg aciddisodium salt dihydrate Heptane 142-82-5 100.21 200 kg Nitrogen Asneeded Pipazoline N/A 531.0 1.0 45.2 24.0 kg Potassium acetate 127-08-298.1417 5.0 225.99 22.2 kg Potassium trifluoro(2-fluoro- N/A 233.03 1.20 54.24 12.6 kg 6-hydroxyphenyl)borate 2-Propanol 67-63-0 66.10 N/A850 kg Si-Thiol N/A N/A N/A 13.2 kg Sodium hydroxide 1310-73-2 40.00 6.5kg USP purified water As needed

To a reactor was added degassed dioxane (74.2 kg), tent-butyl(3S)-4-{7-chloro-6-fluoro-(1M)-1-[4-methyl-2-(propan-2-yl)pyridin-3-yl]-2-oxo-1,2-dihydropyrido[2,3-d]pyrimidin-4-yl}-3-methylpiperazine-1-carboxylate (Compound 6,Pipazoline) (24.0 kg, 45.2 mol), potassium acetate (22.2 kg, 45.2 mol),and (dppf)PdCl₂ (0.74 kg, 1.01 mol). The reactor was inerted withnitrogen gas. The solution was sparged with nitrogen gas until theoxygen content was <500 mg/L. The reaction was heated to 87.5° C. Asolution of potassium trifluoro(2-fluoro-6-hydroxyphenyl)borate (12.6kg, 54.3 mol) in degassed dioxane (49.4 kg) and degassed water (14.4 kg)with oxygen content <500 mg/L was transferred to the reaction,maintaining an internal temperature of 82.5° C.±7.5° C. The reaction wasadjusted to 87.5° C. ±1.5° C. and stirred for 75 min±15 min. A 1.0 MEDTA solution (47.3 kg) followed by water (40.1 kg) was charged to thereactor while maintaining an internal temperature of 85° C.±5° C. Thereaction was cooled to 20° C.±3° C. over >2 h and then stirred for >16h. The reaction was filtered and the crude solids were rinsed with water(3×120 kg). The solids were rinsed with a mixture of heptane (28.8 kg)and 2-propanol (33.1 kg) and then dried at <50° C. for >10 h. A cleanreactor was loaded with crude solids and dichloromethane (240 kg). Thecontents were stirred at 20° C.±5° C. for >30 min. To the reactor wasadded Si-Thiol (144 kg) and dichloromethane (14.9 kg). The reaction wasstirred at 20° C.±5° C. for 18 h. The reaction was filtered and rinsedwith dichloromethane (84 kg). The solution was distilled and solventswapped to 2-propanol. The reaction was heated to 60° C.±3° C. andheptane (108 kg) was charged while maintaining a reaction temperature of60° C.±3° C. The reaction was stirred for 45 min and then cooled andstirred at 20° C.±5° C. for 2.5 h. The reaction was filtered and rinsedwith 50% v/v heptane/2-propanol (61.9 kg). The isolated solids weredried at <50° C. for >12 h to afford tent-butyl(3S)-4-{6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-(1M)-1-[4-methyl-2-(propan-2-yl)pyridin-3-yl]-2-oxo-1,2-dihydropyrido[2,3-d]pyrimidin-4-yl}-3-methylpiperazine-1-carboxylate(Compound 7, BIARYL).

Step 8

General Note: All equivalents and volumes are reported in reference toBIARYL 7

MW Equivalents/ Material CAS # (g/mol) Volumes Moles Theoretical BIARYL7 NA 606.67 1.0 equiv. 5.27 2.75 kg TFA 76-05-1 114.02  11 equiv. 49.75.67 kg DCM 74-09-2 84.93   5 vol NA 13.71 L Methanol 67-56-1 32.04   5vol NA 13.71 L Water 7732-18-5 18.02  20 vol NA 54.8 L Potassium584-08-7 138.20  18 equiv. 94.91 11.24 kg Carbonate DCM 74-09-2 84.93  1 vol NA 2.75 L Water 7732-18-5 18.02  10 vol NA 27.5 L Water7732-18-5 18.02  10 vol NA 27.5 L

To a reactor was added tent-butyl(3S)-4-{6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-(1M)-1-[4-methyl-2-(propan-2-yl)pyridin-3-yl]-2-oxo-1,2-dihydropyrido[2,3-d]pyrimidin-4-yl}-3-methylpiperazine-1-carboxylate(Compound 7, BIARYL) (2.75 kg, 5.27 mol), DCM (13.7 L), and TFA (5.67kg, 49.7 mol). The reaction was stirred for 8-16 h at 20±5° C. To asecond reactor was added potassium carbonate (11.24 kg), water (54.8 L),and methanol (13.7 L) to form a homogenous solution. The reactionmixture was added to the potassium carbonate solution over 2 h. Themixture was stirred at 20±5° C. for an additional 12 h. The resultingslurry was filtered and rinsed with water (2×27.5 L). The wet cake wasdried for 24 h to give6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-4-[(2S)-2-methylpiperazin-1-yl]-(1M)-1-[4-methyl-2-(propan-2-yl)pyridin-3-yl]pyrido[2,3-d]pyrimidin-2(1H)-one(Compound 8, DESBOC).

Step 9

General Note: All equivalents and volumes are reported in reference toDes-BOC

MW Equivalents/ Material CAS # (g/mol) Volumes mmol mass volume Des-BOSNA 506.56 1.0 equiv. 308.4 156.25 g Acryloyl 814-68-6 90.51 1.3 equiv.401.0 36.29 g — chloride¹ — NMP 872-50-4 99.13 4 vol NA — 625 mLN-methyl pyrrolidinone² Water 7732-18-5 18.02 20 vol NA 3125 g 3125 mLNa₂HPO4³ 7558-79-4 141.96 4 equiv. 1233.6 175.12 g — Water 7732-18-518.02 20 vol NA 3125 g 3,125 mL ¹acryloyl chloride was added over 7 minson this scale. Avoid over cooling reaction. Colder reaction temperaturesled to slower reaction time leading to higher levels of m/z 1066impurity as the starting material reacts with the product. Ideal temprange is 22-25 C. ²NMP content of dried cake typically 1-2 wt %.³Disodium phosphate in table is as anhydrous basis. Hydrate may be used,adjust mass accordingly to obtain desired mmol.

6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-4-[(2S)-2-methylpiperazin-1-yl]-(1M)-1-[4-methyl-2-(propan-2-yl)pyridin-3-yl]pyrido[2,3-d]pyrimidin-2(1H)-one(Compound 8, DESBOC) (156.25 g) was combined with N-methyl pyrrolidinone(625mL) and stirred at ambient temperature. To the resulting solutionwas added acryloyl chloride (36.29 g; 401.0mmol) while maintaining <30°C. internal temperature. The contents where stirred for 2 h at 25 C. Ina separate reactor a solution of disodium phosphate (175.1 g; 1234mmo1)in DI water (3.1 L) was prepared. The crude product solution was thentransferred to the reactor containing the disodium phosphate solutionover >2 h at 25° C. The slurry was heated to 45° C. midway through theaddition and after complete addition, aged for 2 h at the sametemperature. The mixture was cooled to 25 C and aged for 4 h beforecollecting the solids by vacuum filtration. The solids where washedtwice with water (1.5 L each) and the product dried under nitrogen andvacuum to afford the product6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-(1M)-1-[4-methyl-2-(propan-2-yl)pyridin-3-yl]-4-[(2S)-2-methyl-4-(prop-2-enoyl)piperazin-1-yl]pyrido[2,3-d]pyrimidin-2(1H)-one(crude Compound 9).⁴ ⁴ Results for this lead lot: 154.06 g isolatedmass, 93% wt, 82.9% corrected yield, 18,000ppm NMP

Step 10

General Note: All equivalents and volumes are reported in reference tocrude drug substance

MW Equivalents/ Material CAS # (g/mol) Volumes mmol mass volume CrudeCompound 9 NA 560.60 1.0 equiv. 253.9 142.33 g — Ethanol (200 proof)64-17-5 — 7.5 V — — 1067 mL USP Water — 18.02 1.9 V — — 270 mL Aceticacid 64-19-7 60.05 1.5 equiv. 380.8 22.87 g 21.82 mL WFI Water — 18.0215.5 vol — — 2200 mL Ethanol (for wash) 64-17-5 — 2.5 V — — 356 mL WFIWater (for wash) — — 5.0 V — — 712 mL Compound 9 seed⁵ — 560.60 0 —0.3-0.7 g — ⁵Seed performs best when reduced in particle size viamilling or with other type of mechanical grinding if mill is notavailable (mortar/pestle). Actual seed utilized will be based on seedavailability. 0.25%-0.5% is seed is target amount.

6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-(1M)-1-[4-methyl-2-(propan-2-yl)pyridin-3-yl]-4-[(2S)-2-methyl-4-(prop-2-enoyl)piperazin-1-yl]pyrido[2,3-d]pyrimidin-2(1H)-one(crude Compound 9) (142.33 g; 253.9mmo1) was combined with ethanol(996mL) and water (270mL). Acetic acid (21.8m1; 380.8mmo1) was added andthe mixture heated to 75° C. to form a solution which was polishfiltered into a clean reactor. The solution was cool to 45° C. and thenwater (1067mL) was added while maintaining an internal temperature >40°C. The solution was seeded with authentic Compound 9 and the resultingmixture aged for 30 min. Water (1138mL) was then added over 2 h. Themixture was cooled to 25° C. and aged for 8 h after which the solid wascollected by vacuum filtration and washed using a mixture of ethanol(355.8mL) and water (711.6mL). The solid was dried using vacuum andnitrogen to obtain6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-(1M)-1-[4-methyl-2-(propan-2-yl)pyridin-3-yl]-4-[(2S)-2-methyl-4-(prop-2-enoyl)piperazin-1-yl]pyrido[2,3-d]pyrimidin-2(1H)-one(Compound 9).

Step A1 Reaction Scheme and Charge Table

MW Equivalents/ Material CAS # (g/mol) Volumes mol Mass (g) Volume (L)3-Fluoroanisole 456-49-5 126.13 1.0 1.19 150 0.136 n-butyllithium (2.5109-72-8 64.06 1.5 1.78 N/A 0.712 M in hexane) diisopropylamine 108-18-9101.19 1.4 1.66 168 0.233 Triethylborate 150-46-9 145.99 2.0 2.38 347.50.405 Tetrahydrofuran 109-99-9 72.11 12 vol N/A N/A 1.8 Hydrochloricacid (2N) 7647-01-0 36.46 10 vol N/A N/A 1.5 Methyl tert-butyl ether1634-04-4 88.15 12 Vol N/A N/A 1.8 Heptane 142-82-5 100.20 10.5 Vol N/AN/A 1.575Reactor A was charged with THF (6 vol) and Diisopropylamine (1.4 equiv).The resulting solution was cooled to −70° C. and n-BuLi (2.5 M inhexane, 1.5 equiv) was slowly added. After addition is complete, asolution of 3-fluoroanisole (1.0 equiv) in THF (6 vol) was added slowlyand kept at −70° C. for 5 min. B(EtO)3 (2.0 equiv) was added slowly andkept at −70° C. for 10 min. The reaction mixture was quenched with 2NHCl. The quenched reaction mixture was extracted with MTBE (3×4 vol).The combined organic phases were concentrated to 1.5-3 total volumes.Heptane (7-9 vol) was added drop-wise and the mixture was cooled to0-10° C. and stirred for 3 h. The mixture was filtrated and rinsed withheptane (1.5 vol). The solid was dried under nitrogen at <30° C. toafford (2-fluoro-6-methoxyphenyl)boronic acid.

Step A2 Reaction Scheme and Charge Table

MW Equivalents/ Material CAS # (g/mol) Volumes mol Mass (g) Volume (L)(2-fluoro-6- 78495-63-3 169.95 1.0 0.118 20 N/A methoxyphenyl) boronicacid. Boron tribromide 10294-33-4 250.52 1.5 0.177 44.2 0.017Dichloromethane 75-09-2 84.93 4 vol N/A N/A 0.080 Water 7732-18-5 18.0213 vol N/A N/A 0.26 Methyl tert-butyl ether 1634-04-4 88.15 13 Vol N/AN/A 0.26 Heptane 142-82-5 100.20 10 Vol N/A N/A 0.20Reactor A was charged with dichloromethane (4 vol) and2-fluoro-6-methoxy-4-methylphenylboronic acid (1 equiv). The reactionmixture was cooled to −30° C. and 1.5 BBr3 (1.5 equiv) was addeddropwise. When the addition completed, the mixture was warmed to 25° C.and stirred 2 h. The reaction mixture was quenched into ice cold (0-5°C.) water (10 vol). MTBE (10 vol) was added and the mixture warmed to25° C. and stirred for 1-2 h or until all solids dissolved. The aqueousphase was separated and extracted with MTBE (3 vol). The combinedorganic extracts were washed with water (3 vol) and then concentrated to1 total volumes. Heptane (10 vol) was added to the mixture and stirredfor 2 h. The resulting product was isolated by filtration and dried at<30° C. to afford (2-fluoro-6-hydroxyphenyl)boronic acid.

Step A3 Reaction Scheme and Charge Table

MW Equivalents/ Material CAS # (g/mol) Volumes mol Mass (kg) Volume (L)(2-fluoro-6- 1256345-60-4 155.92 1.0  89.79 14.00 N/A hydroxyphenyl)boronic acid Citric acid 5949-29-1 210.14 1.64 147.26 30.94 N/Amonohydrate Acetonitrile 75-05-8 41.05 21 Vol N/A 220.1 294 Potassiumfluoride 7789-23-3 58.10 4.00 359.16 20.87 N/A USP Water 7732-18-5 18.022.0 Vol N/A 28.00 28.00 Celite N/A N/A N/A N/A 7.00 N/A 2-Propanol67-63-0 60.10 25 Vol N/A 275 350

Step A3

Potassium Fluoride (21.0 kg; 20.87 mol) was combined with water (28 L)in a reactor (reactor A) and the contents stirred for 30 min. In aseparate reactor (reactor B), (2-fluoro-6-hydroxyphenyl)boronic acid(14.00 kg, 89.79 mol) was charged followed by acetonitrile (206.1 kg)and citric acid (30.94 kg; 147.26 mol) at 25 C. The contents of reactorA was added to reactor B at 25C and stirred at that temperature for 10h. The reaction mixture was filtered through a bed of celite (7.0 kg)and rinsed with acetonitrile (42 kg). The filtrate was combined withisopropanol (56 kg) and then distilled under vacuum at a temperature<35° C. replacing the distilled volume to the reactor with isopropanoland repeated as needed to complete the solvent swap from acetonitrile toisopropanol. The slurry was cooled to 15C and aged for 1 h beforefiltered and washing with 28 kg of isopropanol. The cake was dried usingvacuum and nitrogen and packaged to afford Compound A3.

Resolution of the M-Dione Compound 5

Chromatographic Resolution of M-dione intermediate

Numerous chiral chromatographic techniques and methods were used toisolate the M-dione from Compound 4. The techniques and stationaryphases are well known in the art and are outlined in Table 1.

TABLE 1 Technique Stationary Phase Mobile Phase Yield^(∧) SFCChiralpak ® AD 40% methanol/60% ~95% CO2 * HPLC Chiralpak ® AD 90/10/0.1ethanol/ ~94% methanol/ triethylamine HPLC Chiralpak ® IG 60/40/0.1ethanol/ ~92% methanol/ Triethylamine Simulated Moving Chiralpak ® ICAcetonitrile ~96% Bed (SMB) ^(∧)Yield is defined as % of availableM-Dione that was recovered at the required purity of >98% ee. * Thisseparation was performed multiple times. For each lot of material, themobile phase may have been slightly modified to accommodate forvariations in the lots. Additional mobile phases used for purificationincluded: 1) 25/75 methanol/CO₂, 2) 30/70 methanol/CO₂, and 3) 50/50methanol/CO₂. The SFC, HPLC, and SMB techniques are well known in theart and the Chiralpak ® stationary phases are commercially availablefrom commercial sources, such as Fisher Scientific and DaicelCorporation.

However, it is desired to develop a more efficient process to isolatethe M-Dione (Compound 5).

Classical Resolution

The present invention is directed to the development of a viableclassical resolution process for M/P-Dione racemate (Compound 4).

A total of 100 cocrystal screening experiments were performed and threepotential cocrystals of Dione were identified. Based on the highest arearatio of M/P-Dione in the residual solid and lowest area ratio in thesupernatant, (+)-2, 3-dibenzoyl-D-tartaric acid (DBTA) was selected asthe chiral reagent for resolution.

According to the results from 100 cocrystal screening experiments and 20more solvent screening, 2-MeTHF/n-heptane was found to provide a betterresolution result than other solvent systems. Based on the solubilityresults of M-Dione cocrystal and P-Dione cocrystal in different ratiosof 2-MeTHF and n-heptane, 2-MeTHF/n-heptane (1.4:1, v/v) was selected asthe optimal solvent composition for resolution.

In order to find out any possible form conversion to Dione racemate orM/P-Dione during crystallization process of chiral resolution, thesolubility of M-Dione cocrystal, P-Dione cocrystal, M+P-Dione cocrystalmixture (1:1, w/w), Dione racemate and DBTA were determined at differenttemperatures in 2-MeTHF/n-heptane (1.4:1, v/v). No form change wasobserved for M-Dione cocrystal and P-Dione cocrystal at differenttemperatures for 7 days. However, Dione racemate Type C was obtainedafter stirring of a mixture of M+P-Dione cocrystal mixture (1:1, w/w) atdifferent temperatures for 7 days. Dione racemate Type D (20 and 30° C.)or Dione racemate Type C (40, 50, 60 and 65° C.) were observed afterstirring of Dione racemate at corresponding temperatures for 7 days. Asolubility of ˜100 mg/mL was observed under all the temperatures forDBTA.

To further optimize the resolution process, the ternary phase diagram ofM/P-Dione cocrystal was drawn based on the equilibrium solubilityresults and no eutectic point was obtained likely because racemate TypeC could crystallize out when both M-Dione cocrystal and P-Dionecocrystal were present. Another ternary phase diagram of M/P-Dione wasdrawn based on the equilibrium solubility results and no eutectic pointwas obtained likely because Dione racemate Type C or Type D couldcrystallize out when both M-Dione and P-Dione were present.

In summary, a chiral reagent (DBTA) and a solvent system((2-MeTHF/n-heptane (1.4:1, v/v)) were identified for resolution ofDione racemate. Small scale crystallization process using the resolvingreagent and solvent system could achieve a yield of 39% and ee purity of99% for M-Dione. In addition, polymorphism of Dione racemate wasobserved and investigated during screening experiments.

2 Screening Experiment 2.1 Cocrystal Screening

A total of 100 cocrystal screening experiments were performed using 20acids and 5 solvent systems (Results summarized in Table 2-1). Ingeneral, Dione racemate and acid in the molar ratio at 1:1 was mixed andstirred at RT for 3 days before isolation for XRPD. Based on the XRPDresults, three potential acids that could form cocrystals of Dioneracemate were identified, including (1S)-(−)-camphanic acid (FIG. 2-2),(+)-2, 3-dibenzoyl-D-tartaric acid (FIG. 2-3) and D-(+)-malic acid (FIG.2-4). Four new freebase crystal forms were also obtained based on XRPDresults, which were assigned as Dione racemate Type B˜E.

As showed in Table 2-2, the supernatant and residual solid of the threepotential cocrystals were further tested by HPLC. The ratio ofM-Dione/P-Dione was measured and summarized in Table 2-2. As a result,DBTA cocrystal showed an area ratio of M-Dione/P-Dione at 0.11 in thesupernatant and 4.4 in the residual solid, which suggested that M-Dioneand P-Dione showed a good resolution after forming cocrystal with DBTA.Thus, DBTA was selected as the chiral reagent for further resolutionoptimization.

TABLE 2-1 Summary of cocrystal screening experiments 2-MeTHF/ MTBE/Solvent H₂O/ACN n-heptane n-heptane Acid Acetone (1:1, v/v) EtOAc (1:1,v/v) (1:1, v/v) blank Type B⁺ Type C⁺ Type C⁺ Type D⁺ Type E⁺ L-Asparticacid Type B Type C Type C + acid Type D Type E (R)-1,4-Benzodioxane-2-Type B Type C Type C Type D Type E + acid carboxylic acid(1S)-(−)-Camphanic acid Type B Type C Cocrystal Cocrystal Cocrystal TypeA* Type B* Type A* (−)-Camphoric acid Type B Type C + acid Type C + acidType D Type E + acid (+)-2,3-Dibenzoyl-D-tartaric Type B Type CAmorphous Cocrystal Cocrystal acid Type A^(#) Type A^(#) D-Glutamic acidType B Type C Type C + acid Type D Type E D-(+)-Malic acid Type B Type CCocrystal Type D Type E Type A^($) (R)-(−)-Mandelic acid Type B Type CType C Type D Type E + acid (−)-Menthyloxyacetic acid Type B Type C TypeC Type D Type E (S)-(+)-α- Type B Type C Type C Type D Type EMethoxyphenylacetic acid (R)-(+)-α-Methoxy-α- Type B Type C Type C TypeD Type E trifluoromethylphenylacetic acid (R)-(−)-5-Oxo-2- Type B Type CType C Type D Type E tetrahydrofurancarboxylic acid (R)-(+)-N-(1- Type BType C Type C Type D Type E + acid Phenylethyl)succinamic acid(S)-(+)-2-Phenylpropionic Type B Type C Type C Type D Type E acidL-Pyroglutamic acid Type B Type C Type C Type D Type E + acidD-(−)-Quinic acid Type B + Type C Type C + acid Type D + acid Type E +acid acid L-(+)-Tartaric acid Type B Type C Type C Type D Type EL-Ascorbic acid Type Type C Type C+30acid Type D Type E B+acidN,N-Bis[(R)-(−)-1- Type B Type C Type C Type D Type E + acidphenylethyl]phthalamic acid (S)-phenylsuccinic acid Type B Type C TypeC + acid Type D Type E + acid ⁺Denotes crystal form of free rac-dione(no co-crystal formed), *(1S)-(−)-Camphanic acid cociystal,^(#)(+)-2,3-Dibenzoyl-D-tartaric acid cociystal, ^(s)D-(+)-Malic acidcociystal

TABLE 2-2 HPLC data summary of three cocrystals Peak area SupernatantSolid Sample P-Dione (area) M-Dione (area) M/P P-Dione (area) M-Dione(area) M/P (1S)-(−)-Camphanic acid cocrystal 9021.4 8274.2 0.9 6418.66360.4 1.0 Type A (810465-06-C3) (1S)-(−)-Camphanic acid cocrystal4673.2 4303.4 0.9 4768.3 4736.9 1.0 Type B (810465-06-D3) DBTA cocrystalType A 17673.1 1858.6 0.11 1180.2 5249.8 4.4 (810465-06-D5) D-(+)-Malicacid cocrystal Type A 11382.6 10696.5 0.9 6443.3 6366.7 1.0(810465-06-C7)

2.2 Solvent Screening

To select a suitable solvent to further resolute M-Dione and P-Dione,the area ratio from HPLC of M/P-Dione was collected in 20 moresolvent/solvent mixtures. As listed in Table 2-3, 2-MeTHF showed thebest resolution with the M-Dione/P-Dione area ratio of 0.7 in thesupernatant and 4.1 in the residual solid. However, 2-MeTHF/n-heptane(1:1, v/v) showed a better resolution result during cocrystal screening(Table 2-2), thus 2-MeTHF/n-heptane was selected for furtheroptimization. The area ratio from HPLC of M/P-Dione was collected indifferent ratios of 2-MeTHF/n-heptane with different acid/base ratios.Results in Table 2-4 showed that higher ratio of acid/FB (2:1 or 1.5:1)in 2-MeTHF/n-heptane (8:1 or 4:1, v/v) is desirable to improve the ratioof M/P-Dione in isolated solids.

The solubility of M-cocrystal and P-cocrystal in different ratios of2-MeTHF/n-heptane was also performed at 5 and 25° C., which weresummarized in Table 2-5. M-cocrystal was provided by client andP-cocrystal was prepared via reverse anti-solvent and anti-solvent (theexperimental details refer to Section 4.3). The solubility result inTable 2-5 showed that the volume ratio of 2-MeTHF/n-heptane at 1.5:1could afford the best resolution at RT. More resolution experiments wereperformed by client, from which a volume ratio of 1.4:1 showed the bestresolution result. Thus, the volume ratio of 2-MeTHF/n-heptane at 1.4:1was selected as the solvent system for resolution.

TABLE 2-3 Solvent screening of Dione racemate DBTA cocrystal(M-Dione/P-Dione area ratio) Solvent Supernatant Solid SolventSupernatant Solid 2-MeTHF 0.7 4.1 DMSO/n-heptane (1:1, v/v) NA NA MTBE0.04 3.5 DMF/n-heptane (1:1, v/v) 0.9 1.0 MeOH 0.9 1.0 Toluene/n-heptane(1:1, v/v) 0.8 1.0 IPA 1.0 1.0 Acetic acid NA NA EtOH/n-heptane 0.9 1.0Formic acid NA NA (1:1, v/v) MIBK 0.9 1.0 DCM NA NA MEK 1.0 1.0 Cumene0.9 1.0 IPAc 0.9 1.0 1-Butanol 0.9 1.0 THF NA NA n-Propanol 0.9 1.0 NMPNA NA 1,3 -Dimethy1-2-imidazolidinone 0.9 1.0 NA: A clear solution wasobtained and no solid was isolated.

TABLE 2-4 Results of acid/base ratio and 2-MeTHF/n-heptane ratioscreening with Dione racemate DBTA cocrystal (M-Dione/P-Dione arearatio) Acid/base ratio 2-MeTHF/n- 2:1 1.5:1 1:1.5 1:2 heptane (v/v) FormL S Form L S Form L S Form L S 8:1 CA 0.4 13.4 CA 0.4 8.9 CA 0.5 7.0 CA0.6 2.6 4:1 CA 0.3 8.7 CA 0.3 6.2 CA 0.4 6.2 CA 0.4 5.7 2:1 CA 0.2 6.0CA 0.2 5.6 CA 0.2 4.7 CA 0.2 5.2 1:2 CA 0.3 1.2 CA 0.1 1.3 CA 0.1 1.4 CA0.1 1.7 1:4 DA 1.4 1.0 CA 0.4 1.0 CA + DA 0.1 1.0 CA 0.1 1.0 1:8 CA + DA0.9 1.0 DA 0.5 1.0 DA 0.3 1.0 DA 0.3 1.0 L: Supernatant, S: Solid, CA:Cocrystal Type A, DA: Dione racemate Type A

TABLE 2-5 2-MeTHF/n-heptane ratio screening of M/P-Dione cocrystalsM-Dione P-Dione cocrystal cocrystal Temperature 2-MeTHF/n- SolubilitySolubility (° C.) heptane (v/v) (mg/mL) XRPD (mg/mL) XRPD 5 1:1 11.3Type A* 38.5 Type A^(#) 1.5:1   17.3 Type A* 60.8 Type A^(#) 2:1 22.2Type A* 66.9 Type A^(#) 3:1 28.6 Type A* 86.6 Type A^(#) 4:1 30.8 TypeA* 82.9 Type A^(#) 6:1 47.1 Type A* 92.1 NA 8:1 55.1 Type A* 90.6 NA 251:1 13.4 Type A* 52.9 Type A^(#) 1.5:1   20.3 Type A* 80.8 Type A^(#)2:1 28.1 Type A* 81.7 Type A^(#) 3:1 39.0 Type A* 87.0 NA 4:1 43.0 TypeA* 86.5 NA 6:1 53.2 Type A* 81.9 NA 8:1 65.0 Type A* 89.2 NA*M-cocrystal Type A ^(#)P-cocrystal Type A2.3 Solubility of Dione DBTA cocrystal, Dione racemate and DBTA

The 7-day equilibrium solubility of M-Dione cocrystal, P-Dionecocrystal, M+P-Dione cocrystal mixture (1:1, w/w) and Dione racematewere set up at different temperatures (20, 30, 40, 50, 60, 65, 75 and80° C.) in 2-MeTHF/n-heptane (1.4:1, v/v). Color change was observed at75 and 80° C. after 5 days suggestive of degradation, so the solubilitywas not collected. No form change was observed upon stirring ofM-cocrystal and P-cocrystal at different temperatures for 7 days (FIG.2-5 and FIG. 2-6). Dione racemate Type C was obtained after stirring ofa mixture of M-Dione cocrystal and P-Dione cocrystal mixture (1:1, w/w)at different temperatures for 7 days (FIG. 2-7). Dione racemate Type D(20 and 30° C.) and Dione racemate Type C (40, 50, 60 and 65° C.) wereobserved after stirring of Dione racemate at different temperature for 7days (FIG. 2-8 and FIG. 2-9).

The 5-day equilibrium solubility of DBTA was set up at differenttemperatures (20, 30, 40, 50, 60 and 65° C.) in 2-MeTHF/n-heptane(1.4:1, v/v). A solubility of ˜100 mg/mL was observed under all thetemperatures. No significant difference was observed with varyingtemperatures (Table 2-7).

TABLE 2-6 Solubility of Dione DBTA cocrystal, mixture of M/P-Dionecocrystal, Dione reaemante in 2-MeTHF/n-heptane (1.4:1, v/v) Temp-Solubility Sample erature (mg/mL) ID Material (° C.) M-Dione P-DioneCrystal Form 1-01-A1 M-Dione 20 13.1 — M-cocrystal Type A 1-01-A2cocrystal 30 15.8 — M-cocrystal Type A 1-01-A3 40 18.4 — M-cocrystalType A 1-01-A4 50 17.2 — M-cocrystal Type A 1-01-A5 60 34.6 —M-cocrystal Type A 1-01-A6 65 35.4 — M-cocrystal Type A 1-01-B1 P-Dione,20 — 39.4 P-cocrystal Type A 1-01-B2 cocrystal 30 — 56.5 P-cocrystalType A 1-01-B3 40 — 55.8 P-cocrystal Type A 1-01-B4 50 — 79.9P-cocrystal Type A 1-01-B5 60 — 113.9 P-cocrystal Type A 1-01-B6 65 —110.0 P-cocrystal Type A 1-01-C1 M + P- 20 7.3 10.4 Dione racemate TypeC 1-01-C2 Dione 30 9.2 16.0 Dione racemate Type C 1-01-C3 cocrystal 409.8 12.2 Dione racemate Type C 1-01-C4 mixture 50 12.1 21.9 Dioneracemate Type C 1-01-C5 60 18.7 26.7 Dione racemate Type C 1-01-C6 6513.4* 18.0* Dione racemate Type C 1-01-D1 Dione 20 18.0 15.2 Dioneracemate Type C 1-01-D2 racemate 30 20.1 17.1 Dione racemate Type C1-01-D3 40 11.5 9.9 Dione racemate Type C 1-01-D4 50 14.2 11.8 Dioneracemate Type C 1-01-D5 60 13.7 11.7 Dione racemate Type C 1-01-D6 6515.3 13.1 Dione racemate Type C

TABLE 2-7 Solubility of DBTA in 2-MeTHF/n-heptane (1.4:1, v/v) Sample IDTemperature (° C.) Solubility (mg/mL) 1-15-A1 20 99.1 1-15-A2 30 100.31-15-A3 40 98.9 1-15-A4 50 88.0 1-15-A5 60 105.0 1-15-A6 65 96.1

2.4 Ternary Phase Diagram

2.4.1 M/P-Dione cocrystals

M-Dione cocrystal and P-Dione cocrystal were weighed as correspondingmass listed in the Table 2-8 and were stirred in 2-MeTHF/n-heptane(1.4:1, v/v) at RT for 72 hours. The ternary phase diagram of M/P-Dionecocrystal was drawn based on 72-hour equilibrium solubility data and noeutectic point was obtained (FIG. 2-10).

TABLE 2-8 Solubility data summary for M/P-Dione cocrystals Weight ofWeight of Supernatant Solid M-cocrystal P-cocrystal [M] [P] M P # (mg)(mg) de (%) mg/mL mg/mL de (%) [M]/[P] mg mg de (%) 1 50.7 0 100.0 22.90.0 100.0 NA 27.8 0.0 100.0 2 0 99.7 −100.0 0.0 81.2 −100.0 NA 0.0 18.5−100.0 3 21.1 83.4 −59.6 5.1 59.4 −84.2 11.680 16.0 24.0 −20.0 4 41 83.5−34.1 5.9 45.0 −76.9 7.658 35.1 38.5 −4.6 5 84 85 −0.6 11.5 23.7 −34.92.070 72.5 61.3 8.4 6 167.7 83.1 33.7 12.2 20.8 −26.2 1.711 155.5 62.342.8 7 49.7 50.4 −0.7 17.5 40.8 −40.0 2.331 32.2 9.6 54.1 8 49.7 25 33.119.9 21.8 −4.6 1.095 29.8 3.2 80.7 9 49.2 12.2 60.3 21.3 9.6 37.9 0.45127.9 2.6 82.9

2.4.2 M/P-Dione

M-Dione and P-Dione were weighed as corresponding mass listed in theTable 2-9 and were stirring in 2-MeTHF/n-heptane (1.4:1, v/v) at RT for5 days. The ternary phase diagram was drawn based on 5-day equilibriumsolubility data in 1.0 mL of 2-MeTHF/n-heptane (1.4:1, v/v) at RT.M-Dione Type A, P-Dione Type A, Dione racemate Type C and Type D wereobserved in the residual solid of solubility samples. No eutectic pointswere obtained in the phase diagram (FIG. 2-11).

TABLE 2-9 Data summary of Ternary Phase Diagram for M/P-Dione Weight ofWeight of Supernatant Solid M-Dione P-Dione [M] [P] M P # (mg) (mg) de(%) mg/mL mg/mL de (%) [M]/[P] mg mg de (%) XRPD 1 99.3 0 100.0 80.1 0100.0 NA 19.2 0.0 100.0 A 2 0 100.6 −100.0 0 77.1 −100.0 NA 0.0 23.5−100.0 A 3 20.2 119.3 −71.0 7.7 50.3 −73.6 0.2 12.5 69.0 −69.2 A + RD 489.8 90.3 −0.3 16.2 17.5 −3.9 0.9 73.6 72.8 0.6 RD 5 121.1 40.7 49.753.0 1.6 94.2 33.4 68.1 39.1 27.0 A + RC 6 90.2 19.5 64.4 52.6 1.4 94.837.6 37.6 18.1 35.0 A + RC 7 41.6 39.9 2.1 9.8 9.0 4.4 1.1 31.8 30.9 1.4RC 8 39.2 120.3 −50.8 7.6 48.1 −72.8 0.2 31.6 72.2 −39.1 A + RD 9 119.120.5 70.6 54.1 1.6 94.4 34.5 65.0 18.9 54.9 A + RC RC: Dione racemateType C; RD: Dione racemate Type D; A: M or P-Dione Type A (the XRPDpatterns of M-Dione Type A and P-Dione Type A were same and were notdistinguished).

3 Solid State Characterization of Crystal Forms

A total of five Dione racemate crystal forms and two cocrystal formswere obtained. All these forms were characterized by XRPD, TGA, DSC, PLMand ¹H NMR and summarized in Table 2-10. The solid statecharacterization data suggested Dione racemate Type A and Type D wereidentified as 2-MeTHF solvates, Type B as an acetone solvate, Type C asan anhydrate, and Type E as a MTBE solvate.

Both M-Dione cocrystal Type A and P-Dione cocrystal Type A were found tobe as 2-MeTHF solvates. All the characterization data are demonstratedin FIG. 3-1 to FIG. 3-22.

TABLE 2-10 Summary of crystal forms Endotherm Sample ID Crystal form(peak, ° C.) TGA (wt %) ¹H NMR (wt %) 5-05-A Dione racemate Type A110.2, 248.6, 213.4*  2.5 (150° C.)  2.4 (2-MeTHF) 1-10-A1 Dioneracemate Type B 113.4, 126.0, 250.9  9.8 (150° C.)  6.2 (acetone)1-01-D5 Dione racemate Type C 251.9  3.0 (150° C.) ND^(&) 1-01-D1 Dioneracemate Type D 120.5, 253.3 15.4 (150° C.) 12.0 (2-MeTHF) 1-10-A4 Dioneracemate Type E 151.6, 158.6, 248.7 14.7 (160° C.)  7.5 (MTBE) 5-17-AM-Dione cocrystal Type A 109.6, 119.2  6.6 (125° C.) 10.6 (2-MeTHF)5-16-A P-Dione cocrystal Type A 88.3, 112.3, 132.8  9.2 (140° C.) 10.6(2-MeTHF) *exothermic peak; ^(&)not detected.

3.1.1 Competitive Slurry of Dione Racemate Forms

Dione racemate Type B˜E were successfully re-produced via slurry ofDione racemate Type A in acetone, H20/ACN (1:1, v/v), 2-MeTHF/n-heptane(1.4:1, v/v) and MTBE/n-heptane (1:1, v/v) at RT, respectively.

Around 5 mg of each Dione racemate forms (Type A˜E) were weighed into anHPLC vial, 0.3 mL of saturated Dione racemate solution in2-MeHTF/n-heptane (1.4:1, v/v) was added into the vial and the mixturewas then stirred at 20, 30, 40, 50, 60 and 65° C. for 5 days.

All the freebase forms converted to Dione racemate Type C viacompetitive slurry in 2-MeHTF/n-heptane (1.4:1, v/v) at targettemperatures, suggesting Dione racemate Type C is the mostthermodynamically stable form in 2-MeHTF/n-heptane (1.4:1, v/v) from 20to 65° C.

TABLE 2-11 Competitive slurry results Starting Experiment TemperatureForm ID Solvent (° C.) Solid Form Dione 1-16-B1 2-MeTHF/ 20 Dioneracemate racemate n-heptane Type C Type 1-16-B2 (1.4:1, v/v) 30 Dioneracemate A~E Type C 1-16-B3 40 Dione racemate Type C 1-16-B4 50 Dioneracemate Type C 1-16-B5 60 Dione racemate Type C 1-16-B6 65 Dioneracemate Type C3.2 Preparation of P-Dione cocrystal

3.2.1 Small Scale

2 g P-dione and 1 g DBTA were dissolved in 18 mL 2-MeTHF at 65° C. toget an almost clear solution. 18 mL heptane was added to this solutionin 1 h. The solution was cooled to 20° C. over 4 h and aged overnight.The solution was evaporated using air blow at RT for about 1 h and ayellowish oily-like paste was obtained. Another 54 mL heptane was addedto the mixture with stirring for 2 h. The suspension was filtered. Thesolid sample was assigned as 810465-16-A.

3.2.2 Large Scale

10 g P-dione and 5 g DBTA were dissolved in 100 mL 2-MeTHF at 65° C. Thesolution was filtered by 0.45 p.m PTFE filter and a clear solution wasobtained. The clear solution was added dropwise to a suspension of 400mL heptane containing ˜1 g seeds (810465-16-A) produced from the firstrun. The suspension was kept stirring at RT for 5 h before isolation.About 10 g of the P-Dione cocrystal (810465-20-A) was produced with ayield of ˜66%.

4 Instruments and Methods 4.1 XRPD

For XRPD analysis, PANalytical X-ray powder diffract meters were used inreflection mode. The XRPD parameters used are listed in Table 4-1.

TABLE 4-1 Parameters for XRPD test Parameters PANalytical PANalyticalPANalytical Model Empyrean X′ Pert³ X′ Pert³ X-Ray Cu, kα, Cu, kα, Cu,kα wavelength Kα1 (Å): Kα1 (Å): Kα1 (Å): 1.540598, 1.540598, 1.540598,Kα2 (Å): Kα2 (Å): Kα2 (Å): 1.544426 1.544426 1.544426 Kα2/Kα1 Kα2/Kα1Kα2/Kα1 intensity intensity intensity ratio: 0.50 ratio: 0.50 ratio:0.50 X-Ray tube 45 kV, 45 kV, 45 kV, setting 40 mA 40 mA 40 mADivergence Automatic ⅛° Fixed ⅛° slit Scan mode Continuous ContinuousContinuous Scan range 3°-40° 3°-40° 3°-40° (°2TH) Scan step 17.8 46.718.9 time (s) Step size  0.0167  0.0263  0.0131 (°2TH) Test Time 5 min30 s 5 min 04 s 4 min 15 s

4.2 TGA and DSC

TGA data were collected using a TA discovery 550, Q500 and Q5000 TGAfrom TA Instruments. DSC was performed using Q500, Q5000 and Discovery2500 DSC from TA Instruments. Detailed parameters used are listed inTable 4-2 4-2.

TABLE 4-2 Parameters for TGA and DSC tests Parameters TGA DSC MethodRamp Ramp Sample pan Aluminum, open Aluminum, crimped TemperatureRT-350° C. 25° C.-300 ° C. Heating rate 10° C./min 10° C./min Purge gasN₂ N₂

4.3 HPLC

Agilent 1100/1260 HPLC was utilized to test solubility, with detailedmethods listed in Table 4-3.

TABLE 4-3 HPLC method for solubility test HPLC Agilent 1100 with DADdetector Column CHIRALPAK IC-3, 4.6 × 100 mm, 3 um Mobile phase A:n-Heptane B: MeOH/EtOH (1:1, v/v) Isocratic elution A:B = 75:25, 60:40Run time 10.0 min Post time  0.0 min Flow rate  1.0 mL/min Injectionvolume   5 μL Detector wavelength UV at 215 nm Column temperature 40° C.Sampler temperature RT Diluent EtOH

TABLE 4-4 HPLC method for solubility test (DBTA) HPLC Agilent 1260 withDAD detector Column Agilent ZORBAX 300SB-C3, 150 × 4.6 mm, 3.5 μm Mobilephase A: 0.05% TFA in H₂O B: 0.05% TFA in ACN Isoctatic elution A:B =65:35 Run time 5.0 min Post time 0.0 min Flow rate 0.6 mL/min Injectionvolume   5 μL Detector wavelength UV at 215 nm Column temperature 40° C.Sampler temperature RT Diluent EtOH

4.4 ¹H NMR

¹H NMR spectrum was collected on Bruker 400M NMR Spectrometer usingDMSO-d6 as solvent.

4.5 PLM

Polarized light microscopic picture was captured on Nikon DS-Fi2 uprightmicroscope at room temperature.

Additional screening of compound 5 with1,3-diphenyl-3-oxopropanesulfonic acid 11b.

Due to the low basicity of the pyridine moiety in compound 5 and thelimited ‘hits’ in terms of the formation of crystalline salts using thestandard screening set, it was chosen to screen racemic compound 4 with1,3-diphenyl-3-oxopropanesulfonic acid 1 lb on a 0.06 mmol scale.

Gram scale resolution of racemic compound 5: A 250 mL round-bottom flaskwas charged with 2.0 g racemic compound 4 (5.7 mmol, 1.0 eq.) in 200 mLEtOH:AcOH (90:10 v:v). After the material had dissolved, 832 mg ofsulfonic acid 11b (2.9 mmol, 0.5 eq.) was added to the solution. Theclear solution was left to stir for 15 hours at a stirring speed of 800rpm. A white precipitate had formed which was isolated from the motherliquor. The isolated salt was suspended in CH₂Cl₂ which was treated witha concentrated aqueous NaHCO₃ solution using a separatory funnel. Theorganic layer was isolated and the basic aqueous layer was extractedwith CH₂Cl₂ (2×). The organic layers were combined and dried overNa₂SO₃. Evaporation of the solvent yielded 415 mg of (M)-5 (96% ee) (seeFIG. 6-1).

The clear mother liquor was evaporated to dryness. The yellow oilymaterial was dissolved in CH₂Cl₂ and treated with a concentrated aqueousNaHCO₃ solution using a separatory funnel. The organic layer wasisolated and the basic aqueous layer was extracted with CH₂Cl₂ (2×). Theorganic layers were combined and dried over Na₂SO₃. Evaporation of thesolvent yielded 1579 mg of 5 (23% ee in (P)-atropisomer; FIG. 6-2).

Polymorph Screening on M-Dione DBTA Cocrystal 5 Characterization ofCrystal Forms of M-Dione DBTA Cocrystal

Polymorph screening experiments for the M-Dione were set up under 100conditions using methods of slurry conversion, slow evaporation, slowcooling, anti-solvent addition, vapor diffusion, temperature cycling,and wet grinding. A total of 17 crystal forms (Type A˜Q) were obtainedfrom the screening. The form relationship is shown in FIG. 4-1. Thedetailed characterization data are provided in Table 5-1 and theoverlays of XRPD patterns are shown in FIG. 5-1. Solid statecharacterization results suggested Type G is a hydrate, while the otherTypes are solvates.

5.1 Instruments and Methods 5.1.1 XRPD

XRPD was performed with a Panalytical X′Pert³ Powder XRPD on a Sizero-background holder. The 20 position was calibrated against aPanalytical Si reference standard disc. The parameters used are listedin Table 5-a.

TABLE 5-a Parameters for XRPD test Parameters Reflection Mode X-Raywavelength Cu, kα Kα1 (Å): 1.540598, Kα2 (Å): 1.544426, Kα2/Kα1intensity ratio: 0.50 X-Ray tube setting 45 kV, 40 mA Divergence slitFixed ⅛° Scan mode Continuous Scan range 3-40 (°2TH) Scan step time [s]18.87 Step size  0.0131 (°2TH) Test Time 4 min 15 s

5.1.2 TGA/DSC

TGA data was collected using a TA Discovery 550 TGA from TA Instrument.DSC was performed using a TA Q2000 DSC from TA Instrument. DSC wascalibrated with Indium reference standard and the TGA was calibratedusing nickel reference standard. Detailed parameters used are listed inTable 5-b.

TABLE 5-b Parameters for TGA and DSC test Parameters TGA DSC Method RampRamp Sample pan Platinum, open Aluminum, crimped Temperature RT-desiredtemperature Heating rate 10° C./min Purge gas N₂

5.2 Polymorph Screening

The solubility of Type A (3-05-A) was estimated at RT. Approximately 2mg solids were added into a 3-mL glass vial. Solvents in Table 5-c werethen added stepwise (50/50/200/700 μL) into the vials until the solidswere dissolved or a total volume of 2 mL was reached. Results summarizedin Table 5-c were used to guide the solvent selection in polymorphscreening.

Polymorph screening experiments were performed using differentcrystallization or solid transition methods. The methods utilized andcrystal types identified are summarized in Table 5-c.

TABLE 5-c Approximate solubility of starting material (6010013-05-A) atRT Solvent Solubility (mg/mL) Solvent Solubility (mg/mL) MTBE S < 3.1Acetone S > 52.0 H₂O 2.4 < S < 8.0 DMF S > 40.0 n-Heptane 3.0 < S < 10.0Anisole S > 40.0 Toluene 2.7 < S < 9.0 Acetic acid S > 40.0 Hexanes 2.9< S < 9.7 THF S > 50.0 IPA 26.0 < S < 52.0 ACN S > 46.0 2-MeTHF 24.0 < S< 48.0 CHCl3 S > 50.0 1,4-Dioxane 25.0 < S < 50.0 EtOAc S > 72.0 n-BuOH23.0 < S < 46.0 DMSO S > 72.0 MIBK 38.0 < S < 76.0 MeOH S > 78.0 BuOAcS > 28.0 EtOH S > 68.0 IPAc S > 32.0 — —

TABLE 5-d Summary of polymorph screening experiments No. of MethodExperiments Crystal Type Slurry at RT/5° C. 37 Type A~G, Type J, Type N,Type N Slow Evaporation 16 Type A, Type C, Type D, Type J, Type K, TypeL, Type N, Type O Slow Cooling 9 Type C, Type J, Type L, Type OAnti-solvent Addition 9 Type A, Type C, Type H and Type I Liquid VaporDiffusion 5 Type L, Type M, Type Q Solid Vapor Diffusion 6 Type A and MTemperature Cycling 7 Type A, Type G, Type O Wet Grinding 10 Type ATotal 99 Type A~Q

5.2.1 Slurry at RT

Slurry experiments were conducted at RT in different solvent systems.About 20 mg of Type A (3-05-A) was suspended in 0.2 mL of solvent in a3-mL glass vial. After the suspension was stirred magnetically for 13days at RT, the remaining solids were isolated for XRPD analysis.Results summarized in Table 5-e indicated that Type A˜D and Type J wereobtained.

TABLE 5-e Summary of slurry experiments at RT Experiment ID Solvent(v:v) Solid Form 3-07-A1 MTBE Type B 3-07-A2 H₂O Type A 3-07-A3n-Heptane Low crystallinity 3-07-A4 Toluene Low crystallinity 3-07-A5Hexanes Type A 3-07-A6* IPA Type A 3-07-A7* 2-MeTHF Type B 3-07-A8*1,4-Dioxane Type J 3-07-A9 n-BuOH Low crystallinity 3-07-A10* MIBK TypeA 3-07-A11 BuOAc Type C 3-07-A12 IPAc Type D 3-07-A13* Acetone Amorphous3-07-A14* DMF Type P 3-07-A15 Anisole Type E 3-07-A16 THF/n-Heptane(1:9)Low crystallinity 3-07-A17 2-MeTHF/n-Heptane(1:9) Type A 3-07-A18IPA/H2O(1:9) Type A 3-07-A19 IPAc/H2O(1:9) Type F 3-07-A20n-BuOH/H2O(1:9) Type A 3-07-A21 n-BuOH/MTBE(1:9) Type A 3-07-A22CHCl3/MTBE(1:9) Type A 3-07-A23* MeOH/H2O Amorphous (937:63, aw = 0.2)3-07-A24* MeOH/H2O Type N (844:156, aw = 0.4) 3-07-A25* MeOH/H2O Type G(693:304, aw = 0.6) 3-07-A26 MeOH/H2O Type G (569:431, aw = 0.8) *Solidobtained via slow evaporation at RT

5.2.2 Slow Evaporation

Slow evaporation experiments were performed under 16 conditions.Briefly, 20 mg of Type A (3-05-A) was dissolved in 0.2˜0.8 mL of solventin a 20-mL glass vial. If no dissolution was achieved, suspensions werefiltered using a PTFE (pore size of 0.2 μM) and the filtrates were usedfor the following steps. The visually clear solutions were covered byParafilm® with 5-10 pinholes and subjected to evaporation at RT. Thesolids were isolated for XRPD analysis. The results summarized in Table5-f indicated that Type A, Type C, Type D, Type J, Type K, Type L, TypeN, Type O were obtained.

TABLE 5-f Summary of slow evaporation experiments Experiment ID Solvent(v:v) Solid Form 3-08-A1 Acetic acid Amorphous 3-08-A2 THF Type J3-08-A3 ACN Amorphous 3-08-A4 CHCl3 Amorphous 3-08-A5 EtOAc Type C3-08-A6 DMSO Amorphous 3-08-A7 MeOH Amorphous 3-08-A8 EtOH Type N3-08-A9 1,4-Dioxane Type J 3-08-A10 n-BuOH Type A 3-08-A11 MIBK Type O3-08-A12 BuOAc Type C 3-08-A13 IPAc Type D 3-08-A14 Acetone Type K3-08-A15 DMF Gel 3-08-A16 2-MeTHF Type L

5.2.3 Slow Cooling

Slow cooling experiments were conducted in 9 solvent systems. About 20mg of Type A (3-05-A) was suspended in 1 mL of solvent in a 3-mL glassvial at RT. The suspension was then heated to 50° C., equilibrated fortwo hour and filtered using a PTFE membrane (pore size of 0.20 μm).Filtrates were slowly cooled down to 5° C. at a rate of 0.1° C./min.Results summarized in Table 5-g indicated Type C, Type G, Type J, Type Land Type O s were observed.

TABLE 5-g Summary of slow cooling experiments Experiment ID Solvent(v:v) Solid Form 3-09-A1* IPA Type O 3-09-A2  2-MeTHF Type L 3-09-A3*1,4-Dioxane Type J 3-09-A4* n-BuOH Type L 3-09-A5* Acetic acid Type G(Low crystallinity) 3-09-A6  THF Type J 3-09-A7* ACN Amorphous 3-09-A8 CHCl3 Low crystallinity 3-09-A9* EtOAc Type C *: Solids obtained fromevaporation at RT.

5.2.4 Anti-solvent Addition

A total of 9 anti-solvent addition experiments were carried out. About20 mg of starting material (3-05-A) was dissolved in 0.2-1.4 mL solventto obtain a clear solution. The solution was magnetically stirredfollowed by addition of 0.2 mL anti-solvent stepwise till precipitateappeared or the total amount of anti-solvent reached 15.0 mL. Theobtained precipitate was isolated for XRPD analysis. Results in Table5-h showed that Type A, Type C, Type H and Type I were obtained.

TABLE 5-h Summary of anti-solvent addition experiments Experiment IDSolvent Anti-solvent Solid Form 3-10-A1* H2O Acetone Type H 3-10-A2  THFAmorphous 3-10-A3* DMSO Type I 3-10-A4* MTBE EtOH Type A 3-10-A5* CHCl3Type A 3-10-A6  EtOAc Type A 3-10-A7* Acetone Type C 3-10-A8* n-heptane2-MeTHF Type A 3-10-A9* IPAc Type C *: Solids obtained from evaporationat RT.

5.2.5 Liquid Vapor Diffusion

Five liquid vapor diffusion experiments were conducted. Approximate 20mg of starting material (3-05-A) was dissolved in appropriate solvent toobtain a clear solution in a 3-mL vial. This solution was then placedinto a 20-mL vial with 3 mL of volatile solvents. The 20-mL vial wassealed with a cap and kept at RT allowing sufficient time for organicvapor to interact with the solution. The precipitates were isolated forXRPD analysis. The results summarized in Table 5-i showed that Type L,Type M and Type Q were generated.

TABLE 5-i Summary of liquid vapor diffusion experiments Experiment IDSolvent Anti-solvent Solid Form 3-11-A1 MIBK n-Heptane Type Q 3-11-A2EtOAc IPA Type M 3-11-A3 THF MTBE Type L 3-11-A4 2-MeTHF n-Heptane TypeL  3-11-A5* DMF Toluene Gel *: Solids were obtained via evaporation atRT.

5.2.6 Solid Vapor Diffusion

Solid vapor diffusion experiments were conducted using 6 differentsolvents. Approximate 10 mg of starting material (3-05-A) was weighedinto a 3-mL vial, which was placed into a 20-mL vial with 2 mL ofvolatile solvent. The 20-mL vial was sealed with a cap and kept at RTfor 7 days allowing solvent vapor to interact with sample. The solidswere tested by XRPD and the results summarized in Table 5-j showed thatType A and Type M were generated.

TABLE 5-j Summary of solid vapor diffusion experiments Experiment IDSolvent Solid Form 3-12-A1 EtOH Type M 3-12-A2 MTBE Type A 3-12-A3 H2OType A 3-12-A4 acetone Amorphous 3-12-A5 2-MeTHF Type A 3-12-A6 IPAcType A

5.2.7 Temperature Cycling

Temperature cycling experiments were conducted in 7 solvent systems.About 20 mg of starting material (3-05-A) was suspended in 1 mL ofsolvent in a 3-mL glass vial at RT. The suspension was then heated to50° C., equilibrated for one hour and filtered using a PTFE membrane(pore size of 0.20 pm). Filtrates were slowly cooled down to 5° C. at arate of 0.2° C./min and then heat to 50° C. at a rate of 1° C./min.Repeat the cycle one more time and then cooling to 5° C. at a rate of0.2° C./min. The samples were stored 5° C. before solids were isolatedand analyzed using XRPD. Results summarized in Table 5-k indicated TypeA, Type G and Type O were observed.

TABLE 5-k Summary of temperature cycling experiments Experiment IDSolvent (v:v) Solid Form 3-13-A1* 2-MeTHF Type A 3-13-A2* MeOH Type G3-13-A3* MIBK Type A 3-13-A4* ACN Amorphous 3-13-A5 CHCl3 Lowcrystallinity 3-13-A6* Toluene Amorphous 3-13-A7* IPA Type O *: Solidsobtained from evaporation at RT.

5.2.8 Slurry at 5° C.

Slurry experiments were conducted at 5° C. in different solvent systems.About 20 mg of starting material (3-05-A) was suspended in 0.2 mL ofsolvent in a 3-mL glass vial. After the suspension was stirredmagnetically for 7 days at 5° C., the remaining solids were isolated forXRPD analysis. Results summarized in Table 5-1 indicated that Type A,Type C Type E and Type J were obtained.

TABLE 5-l Summary of slurry experiments at 5° C. Experiment ID Solvent(v:v) Solid Form 3-14-A1* BuOAc Type C 3-14-A2* IPAc Type D 3-14-A3*Acetone Low crystallinity 3-14-A4* DMF Gel 3-14-A5  Anisole Type E3-14-A6* 2MeTHF Type A 3-14-A7* ACN Low crystallinity 3-14-A8* CHCl3 Lowcrystallinity 3-14-A9* EtOAc Low crystallinity  3-14-A10* MeOH Type C 3-14-A11* THF Type J *: Solids obtained from evaporation at RT.

5.2.9 Wet Grinding

Wet grinding experiments were performed under five conditions. Briefly,10 mg of Type A (3-05-A) was put in mortar and grinding in ˜20 μL ofsolvent for 5 min. The solids were isolated for XRPD analysis. Theresults summarized in Table 5-m indicated that Type A was obtained.

TABLE 5-m Summary of wet grinding experiments Experiment ID Solvent(v:v) Solid Form 3-15-A1 MTBE Amorphous 3-15-A2 H₂O Amorphous 3-15-A3n-Heptane Amorphous 3-15-A4 Toluene Amorphous 3-15-A5 Hexanes Amorphous3-15-A6 IPA Amorphous 3-15-A7 2-MeTHF Type A (low crystallinity) 3-15-A81,4-Dioxane Type A 3-15-A9 n-BuOH Amorphous  3-15-A10 MIBK Amorphous

TABLE 5-1 Characterization of M-dione DBTA cocrystal crystal formsCrystal Form Preparation Weight Loss DSC Endo. (Batch No.) Conditions inTGA (%) (Peak, ° C.) Form ID Type A Classic 7.31 up to 109.4 2-MeTHF(3-05-A) resolution with 125° C. 120.0 solvate DBTA (2- MeTHF) Type BSlurry at RT 7.21 up to 115.7 MTBE (3-07-A1) (MTBE) 125° C. solvate TypeC Slow 7.99 up to 92.7 EtOAc (3-08-A5) Evaporation 125° C. 116.4 solvate(EtOAc) Type D Slurry at RT 7.51 up to 75.4, 110.5, IPAc (3-07-A12)(IPAc) 130° C. 148.0, 116.6, solvate 265.9(exo). Type E Slurry at RT8.63 up to 103.8 119.0 Anisole (3-07-A15) (Anisole) 125° C. Solvate TypeF Slurry at RT 6.2 up to 86.5, 107.9 IPAc (3-07-A19) IPAc/H₂O 130° C.solvate (v:v 1:9) Type G Slurry at RT 6.44 up to 86.0 Hydrate (3-07-A26)(MeOH/H₂O 100° C. 127.2 133.1 aw = 0.8) Type H Anti-Solvent 3.58 up to107.6 Acetone (3-10-A1) (Acetone/H₂O) 130° C. solvate Type IAnti-Solvent 7.26 up to 128.9 DMSO (3-10-A3) (DMSO/H₂O) 150° C. solvateType J Slow 7.27% by 115.7 THF (3-08-A2) Evaporation 125° C. Solvate(THF) Type K Slow 5.71 up to 96.7, 119.8 Acetone (3-08-A14) Evaporation150° C. 147.4 solvate (Acetone) 157.4° C.(exo). Type L Liquid Vapor 7.94up to 126.2 2-MeTHF (3-11-A4) Diffusion 130° C. solvate (2-MeTHF/n-Heptane) Type M Liquid Vapor 3.73 up to 122.6 IPA (3-11-A2) Diffusion150° C. solvate (EtOAc/IPA) Type N Slow 4.08 up to 85.4, 126.5 EtOH(3-08-A8) Evaporation 150° C. 150.9 solvate (EtOH) Type O Slow 2.03 upto 106.2, 151.2 MIBK (3-08-A11) Evaporation 130° C. solvate (MIBK) TypeP Slow 8.33 up to 89.9 DMF (3-07-A14) Evaporation 130° C. solvate (DMF)Type Q Liquid Vapor 6.00 up to 92.9, 148.9 MIBK (3-11-A1) Diffusion 120°C. 170.0 solvate (MIBK/n- Heptane)

5.3 Type A

Type A (3-05-A) was provided by client. The XRPD result showed in FIG.5-4 suggested crystalline. As shown by TGA and DSC data in FIG. 5-5, aweight loss of 7.3% up to 125° C. and two endotherms at 109.4 and 120.0°C. (peak) were observed. As displayed in FIG. 5-6, the presence of2-MeTHF was evidenced in ¹H NMR spectrum. Based on the results, Type Awas considered as a 2-MeTHF solvate.

5.4 Type B

Type B sample (3-07-A1) was obtained via slurry of Type A in MTBE at RT.XRPD pattern shown in FIG. 5-7 suggested crystalline. As shown by TGAand DSC data in FIG. 5-10, a weight loss of 7.2% up to 125° C. and anendotherm at 115.7° C. (peak) were observed. As displayed in FIG. 5-9,the presence of MTBE was evidenced in ¹H NMR spectrum. Based on theresults, Type B was likely a MTBE solvate.

5.5 Type C

Type C sample (3-08-A5) was obtained via slow evaporation in EtOAc atRT. XRPD pattern shown in FIG. 5-10 suggested crystalline. TGA and DSCdata displayed in FIG. 5-11 indicated a weight loss of 8.0% up to 125°C. and two endotherms at 92.7° C. and 116.4° C. (peak). As displayed inFIG. 5-12, the presence of EtOAc was evidenced in ¹H NMR spectrum. Basedon the results, Type C was likely an EtOAc solvate.

5.6 Type D

Type D (3-07-A12) was obtained by slurry of Type A in IPAc at RT. TheXRPD result showed in FIG. 5-13 suggested crystalline. As shown by TGAand DSC data in FIG. 5-14, a weight loss of 7.5% up to 130° C. andendotherms at 75.4° C., 110.5° C., 148.0° C. and 116.6° C. (peak) and anexotherm at 265.9° C. were observed. As displayed in FIG. 5-15, thepresence of IPAc was evidenced in ¹H NMR spectrum. Based on the results,Type D was likely an IPAc solvate.

5.7 Type E

Type E (3-07-A15) was obtained via slurry of Type A in anisole at RT.The XRPD result showed in FIG. 5-16 suggested crystalline. As shown byTGA and DSC data in FIG. 5-17, a weight loss of 8.6% up to 125° C. andtwo endotherms at 103.8° C. and 119.0° C. (peak) were observed. Asdisplayed in FIG. 5-18, the presence of IPAc was evidenced in ¹H NMRspectrum. Based on the results, Type E was likely an anisole solvate.

5.8 Type F

Type F (3-07-A19) was obtained via slurry of Type A in IPAc/H20 (v:v1:9) at RT. The XRPD result showed in FIG. 5-19 suggested crystalline.As shown by TGA and DSC data in FIG. 5-20, a weight loss of 6.2% up to130° C. and two endotherms at 86.5° C. and 107.9° C. (peak) wereobserved. As displayed in FIG. 5-18, the presence of IPAc was evidencedin ¹H NMR spectrum. Based on the results, Type F was likely an IPAcsolvate.

5.9 Type G

Type G (3-07-A26) was obtained via slurry of Type A in MeOH/H20 (aw=0.8)at RT. The XRPD result showed in FIG. 5-22 suggested crystalline state.As shown by TGA and DSC data in FIG. 5-23, a weight loss of 6.4% up to100° C. and endotherms at 86.0° C., 127.2° C. and 133.1° C. (peak) wereobserved. As displayed in FIG. 5-24, no signal for MeOH or MeTHF wasobserved in solution ¹HNMR spectrum. Based on the results, Type G waslikely a hydrate.

5.10 Type H

Type H (3-10-A1) was obtained via anti-solvent addition usingAcetone/H₂O. The XRPD result showed in FIG. 5-25 suggested crystalline.As shown by TGA and DSC data in FIG. 5-26, a weight loss of 3.6% up to130° C. and an endotherm at 107.6° C. (peak) were observed. As displayedin FIG. 5-27, the presence of acetone was evidenced in ¹H NMR spectrum.Based on the results, Type H was likely an acetone solvate.

5.11 Type I

Type I (3-10-A3) was obtained via anti-solvent addition using DMSO/H₂O.The XRPD result showed in FIG. 5-28 suggested crystalline. As shown byTGA and DSC data in FIG. 5-29, a weight loss of 7.3% up to 150° C. andan endotherm at 128.9° C. (peak) were observed. As displayed in FIG.5-30, the presence of DMSO was evidenced in ¹H NMR spectrum. Based onthe results, Type I was likely a DMSO solvate.

5.12 Type J

Type J (3-08-A2) was obtained via slow evaporation in THF. The XRPDresult showed in FIG. 5-31 suggested crystalline. As shown by TGA andDSC data in FIG. 5-32, a weight loss of 7.3% up to 125° C. and anendotherm at 115.7° C. (peak) were observed. As displayed in FIG. 5-33,the presence of THF was evidenced in ¹H NMR spectrum. Based on theresults, Type J was likely a THF solvate.

5.13 Type K

Type K (3-08-A14) was obtained via slow evaporation in acetone. The XRPDresult showed in FIG. 5-34 suggested crystalline. As shown by TGA andDSC data in FIG. 5-35, a weight loss of 5.7% up to 150° C. andendotherms at 96.7° C., 119.8° C. and 147.4° C. (peak) and an exothermat 157.4° C. (peak) were observed. As displayed in FIG. 5-36, thepresence of acetone was evidenced in ¹H NMR spectrum. Based on theresults, Type K was likely an acetone solvate.

5.14 Type L

Type L (3-11-A4) was obtained via liquid vapor diffusion in2-MeTHF/n-Heptane. The XRPD result showed in FIG. 5-37 suggestedcrystalline with preferred orientation. As shown by TGA and DSC data inFIG. 5-38, a weight loss of 7.9% up to 130° C. and an endotherm at126.2° C. (peak) were observed. As displayed in FIG. 5-39, the presenceof acetone was evidenced while no signal for n-heptane was observed in¹H NMR spectrum. Based on the results, Type L was likely a 2-MeTHFsolvate.

5.15 Type M

Type M (3-11-A2) was obtained from liquid vapor diffusion in EtOAc/IPA.The XRPD result showed in FIG. 5-40 suggested crystalline. As shown byTGA and DSC data in FIG. 5-41, a weight loss of 3.7% up to 150° C. andan endotherm at 122.6° C. (peak) were observed. As displayed in FIG.5-42, the presence of IPA was evidenced while no signal for EtOAc wasobserved in ¹H NMR. Based on the results, Type M was likely an IPAsolvate.

5.16 Type N

Type N (3-08-A8) was obtained via slow evaporation in EtOH. The XRPDresult showed in FIG. 5-43 suggested crystalline. As shown by TGA andDSC data in FIG. 5-44, a weight loss of 4.1% up to 150° C. andendotherms at 85.4° C., 126.5° C. and 150.9° C. (peak) were observed. Asdisplayed in FIG. 5-45, the presence of EtOH was evidenced in ¹H NMRspectrum. Based on the results, Type N was likely an EtOH solvate.

5.17 Type O

Type O (3-08-A11) was obtained via slow evaporation in MIBK. The XRPDresult showed in FIG. 5-46 suggested crystalline. As shown by TGA andDSC data in FIG. 5-47, a weight loss of 2.0% up to 130° C. and twoendotherms at 106.2° C. and 151.2° C. (peak) were observed. As displayedin FIG. 5-48, the presence of MIBK was evidenced in ¹H NMR spectrum.Based on the results, Type O was likely a MIBK solvate.

5.18 Type P

Type P (3-07-A14) was obtained via slow evaporation in DMF. The XRPDresult showed in FIG. 5-49 suggested crystalline. As shown by TGA andDSC data in FIG. 5-50, a weight loss of 8.3% up to 130° C. and anendotherm at 89.9° C. (peak) were observed. As displayed in FIG. 5-51,the presence of DMF was evidenced in ¹H NMR spectrum. Based on theresults, Type P was likely a DMF solvate.

5.19 Type Q

Type Q (3-11-A1) was obtained via liquid vapor diffusion inMIBK/n-Heptane. The XRPD result showed in FIG. 5-52 suggestedcrystalline. As shown by TGA and DSC data in FIG. 5-53, a weight loss of6.0% up to 120° C. and endotherms at 92.9° C., 148.9° C. and 170.0° C.(peak) were observed. As displayed in FIG. 5-54, the presence of MIBKwas evidenced while no signal of N-Heptane observed in ¹H NMR spectrum.Based on the results, Type Q was likely a MIBK solvate.

6. Crystal Data and Experimental for Composition 4a

Experimental. Single colourless plate-shaped crystals of (Composition4a) were used as received. A suitable crystal (0.28×0.18×0.09) mm³ wasselected and mounted on a nylon loop with paratone oil on a BrukerAPEX-II CCD diffractometer. The crystal was kept at T=173(2) K duringdata collection. Using Olex2 (Dolomanov et al., 2009), the structure wassolved with the XT (Sheldrick, 2015) structure solution program, usingthe Intrinsic Phasing solution method. The model was refined withversion of XL (Sheldrick, 2008) using Least Squares minimisation.

Crystal Data. C₆₅H₇₂Cl₂F₂N₈O₁₅, MY=1314.20, triclinic, P1 (No. 1),a=11.5683(10) Å, b=11.6705(10) Å, c=13.9593(12) Å, a=68.1780(10)°,β=69.4150(10)°, γ=87.7760(10)°, V=1628.7(2) Å³, T=173(2) K, Z=1, Z′=1,μ(MoK_(α))=0.178, 26758 reflections measured, 11949 unique(R_(int)=0.0528) which were used in all calculations. The final wR₂ was0.2465 (all data) and R₁ was 0.0835 (I>2(I)).p

TABLE 6-2 Fractional Atomic Coordinates (×10⁴) and Equivalent IsotropicDisplacement Parameters (Å² × 10³) for COMPOSITION 4A. U_(eq) is definedas ⅓ of the trace of the orthogonalised U_(ij). Atom x y z U_(eq) O1C5607(5) 3451(5) 6493(5) 44.9(14) O2C 6723(6) 4580(6) 6921(5) 49.9(15)O3C 3704(5) 3216(6) 5826(5) 45.6(14) O4C 2800(6) 3841(6) 4560(6)55.9(16) O5C 3369(5) 5630(6) 6567(5) 50.2(15) O6C 3636(5) 3990(6)7917(5) 50.8(15) O7C 5703(6) 2984(7) 4257(5) 55.4(16) O8C 6595(5)4867(6) 3832(5) 49.1(15) C1C 5032(7) 4527(8) 6031(7) 41(2) C2C 4615(7)4267(8) 5203(7) 39.1(19) C3C 3928(8) 4667(8) 6968(7) 42(2) C4C 5705(8)3954(9) 4361(7) 42(2) CSC 6393(8) 3601(8) 6961(7) 42(2) C6C 6820(9)2403(8) 7493(8) 49(2) C7C 7721(15) 2378(12) 7941(13) 98(3) C8C 8179(15)1311(13) 8410(13) 94(3) C9C 7697(17) 223(14) 8497(14) 107(5) C10C6708(16) 185(12) 8178(13) 98(3) C11C 6265(15) 1303(12) 7665(13) 94(3)C12C 2953(8) 3028(9) 5341(7) 44(2) C13C 2359(9) 1762(9) 5860(8) 54(2)C14C 1223(9) 1501(10) 5822(8) 56(2) C15C 671(11) 300(12) 6312(10) 75(3)C16C 1261(14) −658(13) 6805(13) 98(5) C17C 2446(14) −407(13) 6720(13)97(4) C18C 2954(13) 800(12) 6325(12) 87(4) CUB 6935(2) 1601(2) 11106(2)66.0(7) F1B 4643(5) 1859(6) 10647(5) 66.7(16) O1B 8509(6) 7968(6)7303(6) 58.0(17) O2B 4575(6) 6378(7) 8234(6) 66.1(19) N1B 7896(6)6005(6) 8607(6) 40.4(16) N2B 7345(6) 3931(7) 9798(6) 42.2(17) N3B6567(7) 7139(7) 7745(6) 49.2(18) N4B 11242(6) 5811(7) 8040(6) 47.4(18)C1B 7710(8) 7096(9) 7832(8) 48(2) C2B 7030(8) 4974(8) 9152(6) 40(2) C3B6537(8) 2952(9) 10275(7) 47(2) C4B 5411(8) 2909(8) 10151(7) 47(2) CSB5070(8) 3977(10) 9543(7) 49(2) C6B 5884(8) 5054(9) 9031(7) 44(2) C7B5584(8) 6210(9) 8328(8) 50(2) C8B 9103(7) 5996(7) 8717(7) 41(2) C9B9205(8) 6331(8) 9550(7) 43(2) C10B 10370(9) 6393(9) 9588(7) 49(2) C11B11350(8) 6131(8) 8840(7) 44(2) C12B 10113(7) 5734(7) 7981(7) 38.9(19)C13B 8078(9) 6634(9) 10360(8) 55(2) C14B 10040(9) 5292(9) 7127(8) 54(2)C15B 10546(10) 4041(10) 7234(10) 65(3) C16B 10688(11) 6301(12) 5939(9)73(3) C11A 11316(3) −588(3) 4003(3) 82.7(9) F1A 13799(6) −251(6) 4033(5)77.9(18) O1A 12158(6) 6025(6) 1115(6) 57.3(17) O2A 15430(6) 4363(7)1981(6) 63.4(18) N1A 11949(6) 3944(6) 2039(6) 38.9(16) N2A 11735(7)1804(7) 3004(6) 48.6(19) N3A 13776(6) 5170(7) 1580(6) 45.5(18) N4A8608(6) 4153(7) 2689(6) 46.4(18) CIA 12607(8) 5110(9) 1551(7) 47(2) C2A12449(7) 2876(8) 2555(7) 40(2) C3A 12216(10) 797(10) 3464(8) 56(2) C4A13389(10) 813(9) 3546(8) 56(3) CSA 14083(9) 1886(10) 3114(8) 54(2) C6A13638(8) 2995(8) 2564(7) 44(2) C7A 14365(8) 4199(9) 2043(8) 48(2) C8A10709(7) 3834(8) 2039(6) 37.4(19) C9A 10540(8) 3371(9) 1305(8) 47(2)C10A 9326(8) 3313(8) 1314(7) 47(2) C11A 8421(8) 3689(8) 2008(7) 45(2)C12A 9733(7) 4227(7) 2738(7) 39.4(19) C13A 11568(8) 2995(10) 522(8)55(2) C14A 9890(8) 4703(9) 3549(7) 50(2) C15A 9619(11) 6056(11) 3255(11)71(3) C16A 9045(10) 3875(11) 4734(8) 64(3) O3S 355(10) 1070(10) 9718(9)116(3) C11S −300(40) −220(40) 11630(40) 142(14) C11T −1050(40) −700(50)11410(40) 190(20) C12S 160(30) −310(30) 10410(20) 180(9) C13S 1360(40)−510(50) 10600(40) 151(11) C13T 1220(40) −820(40) 9990(40) 151(11) C14S2240(20) 280(20) 9240(20) 155(8) C15S 1510(20) 1150(20) 8834(18) 139(7)O2S 6461(8) −721(8) 5995(7) 83(2) C6S 5780(20) −1920(20) 5275(19) 145(7)C7S 6000(20) −720(20) 5170(20) 155(8) C8S 7200(30) −180(30) 4150(30)241(15) C9S 7990(20) 670(20) 4390(20) 153(7) C10S 7490(30) 340(30)5490(30) 202(11) O1S 4966(9) 7468(9) 1097(8) 99(3) C1S 5930(20) 8160(20)2110(20) 176(9) C2S 5018(15) 7306(16) 2147(14) 105(5) C3S 3770(20)7280(20) 2920(20) 146(7) C4S 3200(30) 8270(30) 2170(30) 207(12) C5S4180(20) 8430(20) 990(20) 162(8)

TABLE 6-3 Anisotropic Displacement Parameters (× 10⁴) COMPOSITION 4A.Atom U₁₁ U₂₂ U₃₃ U₂₃ U₁₃ U₁₂ O1C 37 (3) 57 (4) 52 (3) −27 (3) −25 (3) 9(3) O2C 47 (4) 54 (4) 52 (4) −21 (3) −21 (3) 4 (3) O3C 33 (3) 63 (4) 39(3) −21 (3) −10 (3) −3 (3) O4C 52 (4) 68 (4) 53 (4) −20 (4) −26 (3) 1(3) O5C 34 (3) 56 (4) 53 (4) −19 (3) −11 (3) 9 (3) O6C 44 (3) 71 (4) 35(4) −21 (3) −11 (3) 3 (3) O7C 46 (4) 71 (5) 56 (4) −34 (4) −15 (3) 3 (3)O8C 33 (3) 56 (4) 55 (4) −22 (3) −11 (3) 7 (3) C1C 31 (4) 48 (5) 47 (5)−20 (4) −14 (4) 1 (4) C2C 35 (5) 42 (5) 41 (5) −16 (4) −14 (4) 2 (4) C3C37 (5) 52 (5) 39 (5) −21 (4) −14 (4) 2 (4) C4C 36 (5) 51 (5) 45 (5) −17(4) −22 (4) 6 (4) C5C 40 (5) 48 (5) 35 (4) −16 (4) −11 (4) 5 (4) C6C 58(6) 45 (5) 51 (5) −22 (4) −26 (5) 10 (4) C7C 141 (9) 68 (6) 117 (8) −37(6) −85 (8) 23 (6) C8C 124 (9) 77 (6) 120 (8) −41 (6) −86 (7) 41 (6) C9C155 (14) 87 (10) 133 (13) −53 (9) −107 (12) 68 (10) C10C 141 (9) 68 (6)117 (8) −37 (6) −85 (8) 23 (6) C11C 124 (9) 77 (6) 120 (8) −41 (6) −86(7) 41 (6) C12C 35 (5) 65 (6) 38 (5) −26 (5) −15 (4) 3 (4) C13C 50 (6)59 (6) 56 (6) −23 (5) −23 (5) 6 (5) C14C 47 (6) 71 (7) 58 (6) −31 (5)−22 (5) 1 (5) C15C 54 (6) 101 (10) 73 (8) −34 (7) −22 (6) −21 (6) C16C96 (10) 81 (9) 114 (11) −3 (8) −67 (9) −27 (8) C17C 98 (10) 73 (8) 110(11) −10 (8) −51 (9) −2 (7) C18C 88 (9) 77 (8) 102 (10) −20 (7) −56 (8)−9 (7) Cl1B 67.9 (16) 57.4 (14) 64.2 (16) −10.1 (12) −27.3 (13) −6.4(12) F1B 67 (4) 76 (4) 55 (3) −19 (3) −22 (3) −23 (3) O1B 41 (4) 55 (4)71 (4) −12 (3) −23 (3) −6 (3) O2B 36 (4) 89 (5) 78 (5) −31 (4) −28 (3)11 (3) N1B 23 (3) 42 (4) 51 (4) −12 (3) −13 (3) −6 (3) N2B 33 (4) 54 (5)35 (4) −14 (3) −10 (3) −5 (3) N3B 40 (4) 58 (5) 48 (4) −12 (4) −23 (4) 2(4) N4B 35 (4) 47 (4) 59 (5) −17 (4) −18 (4) 7 (3) C1B 38 (5) 55 (6) 58(6) −27 (5) −22 (4) 9 (5) C2B 37 (5) 53 (5) 27 (4) −15 (4) −9 (4) 1 (4)C3B 46 (5) 55 (6) 40 (5) −19 (4) −14 (4) 6 (4) C4B 49 (5) 54 (6) 36 (5)−16 (4) −14 (4) −12 (4) C5B 29 (5) 83 (7) 35 (5) −25 (5) −9 (4) −6 (5)C6B 32 (5) 64 (6) 37 (5) −21 (4) −10 (4) −8 (4) C7B 38 (5) 67 (6) 50 (5)−23 (5) −19 (4) 3 (4) C8B 34 (5) 43 (5) 48 (5) −15 (4) −19 (4) 2 (4) C9B39 (5) 51 (5) 42 (5) −22 (4) −12 (4) 1 (4) C10B 49 (6) 59 (6) 42 (5) −20(4) −20 (4) 10 (4) C11B 42 (5) 50 (5) 46 (5) −17 (4) −23 (4) 4 (4) C12B28 (4) 42 (5) 47 (5) −17 (4) −14 (4) 2 (3) C13B 44 (5) 62 (6) 52 (6) −25(5) −8 (4) 7 (5) C14B 40 (5) 62 (6) 61 (6) −26 (5) −18 (5) 0 (4) C15B 52(6) 72 (7) 89 (8) −52 (6) −22 (6) 7 (5) C16B 66 (7) 99 (9) 50 (6) −23(6) −21 (5) −10 (6) Cl1A 84 (2) 60.8 (16) 85 (2) −4.6 (15) −34.1 (16)−3.2 (14) F1A 89 (5) 73 (4) 76 (4) −21 (3) −45 (4) 31 (3) O1A 45 (4) 60(4) 64 (4) −17 (4) −23 (3) 1 (3) O2A 42 (4) 87 (5) 76 (5) −38 (4) −33(3) 8 (3) N1A 26 (3) 53 (4) 37 (4) −16 (3) −12 (3) 1 (3) N2A 41 (4) 59(5) 37 (4) −13 (4) −9 (3) 0 (4) N3A 31 (4) 58 (4) 51 (4) −27 (4) −12 (3)−4 (3) N4A 36 (4) 59 (5) 40 (4) −16 (4) −12 (3) 1 (3) C1A 44 (5) 57 (6)41 (5) −21 (4) −16 (4) 2 (5) C2A 30 (4) 65 (6) 33 (4) −26 (4) −13 (4) 12(4) C3A 61 (6) 65 (6) 48 (6) −20 (5) −28 (5) 6 (5) C4A 57 (6) 55 (6) 60(6) −29 (5) −19 (5) 18 (5) C5A 54 (6) 79 (7) 48 (5) −36 (5) −27 (5) 18(5) C6A 34 (5) 59 (6) 45 (5) −26 (4) −15 (4) 13 (4) C7A 35 (5) 72 (6) 47(5) −34 (5) −13 (4) 5 (5) C8A 32 (4) 46 (5) 30 (4) −10 (4) −12 (4) 6 (4)C9A 33 (5) 61 (6) 48 (5) −18 (4) −16 (4) 5 (4) C10A 38 (5) 59 (6) 46 (5)−17 (4) −18 (4) −5 (4) C11A 30 (5) 59 (6) 38 (5) −14 (4) −8 (4) −1 (4)C12A 31 (4) 47 (5) 34 (4) −9 (4) −10 (4) 3 (4) C13A 39 (5) 91 (7) 41 (5)−34 (5) −10 (4) 12 (5) C14A 36 (5) 73 (6) 46 (5) −31 (5) −12 (4) 5 (4)C15A 64 (7) 81 (8) 78 (8) −44 (6) −22 (6) 6 (6) C16A 58 (6) 96 (8) 39(5) −21 (5) −25 (5) 16 (6) O3S 113 (8) 120 (8) 98 (7) −25 (6) −37 (6) 19(7) The anisotropic displacement factor exponent takes the form:−2π²[h²a*² × U₁₁ + . . . + 2hka* × b* × U₁₂]$

TABLE 6-3 Bond Lengths in Å for COMPOSITION 4A. Atom Atom Length/Å O1CC1C 1.439(10) O1C C5C 1.341(10) O2C C5C 1.194(10) O3C C2C 1.427(10) O3CC12C 1.341(10) O4C C12C 1.215(11) O5C C3C 1.313(10) O6C C3C 1.193(10)O7C C4C 1.194(10) O8C C4C 1.306(10) C1C C2C 1.525(11) C1C C3C 1.528(12)C2C C4C 1.532(12) C5C C6C 1.478(13) C6C C7C 1.386(16) C6C C11C 1.358(16)C7C C8C 1.358(17) C8C C9C 1.35(2) N3B C1B 1.367(11) N3B C7B 1.385(12)N4B C11B 1.346(11) N4B C12B 1.346(10) C2B C6B 1.391(12) C3B C4B1.378(13) C4B C5B 1.357(13) C5B C6B 1.389(12) C6B C7B 1.456(14) C8B C9B1.400(12) C8B C12B 1.369(12) C9B C10B 1.372(12) C9B C13B 1.526(13) C10BC11B 1.359(13) C12B C14B 1.492(13) C14B C15B 1.526(14) C14B C16B1.562(15) C11A C3A 1.714(11) F1A C4A 1.336(11) O1A CIA 1.216(11) O2A C7A1.223(11) N1A C1A 1.386(11) N1A C2A 1.405(11) N1A C8A 1.446(10) N2A C2A1.331(11) N2A C3A 1.321(12) N3A C1A 1.373(12) N3A C7A 1.368(12) N4A C11A1.335(11) N4A C12A 1.335(11) C2A C6A 1.394(12) C3S C4S 1.53(3) C4S C5S1.59(4) C9C C10C 1.373(19) C10C C11C 1.411(17) C12C C13C 1.454(14) C13CC14C 1.384(13) C13C C18C 1.366(16) C14C C15C 1.375(15) C15C C16C1.375(18) C16C C17C 1.368(19) C17C C18C 1.375(18) CUB C3B 1.732(10) F1BC4B 1.339(10) O1B C1B 1.212(11) O2B C7B 1.221(11) N1B C1B 1.398(12) N1BC2B 1.381(10) N1B C8B 1.458(10) N2B C2B 1.344(11) N2B C3B 1.305(11) C3AC4A 1.402(14) C4A C5A 1.324(14) C5A C6A 1.419(13) C6A C7A 1.453(13) C8AC9A 1.393(12) C8A C12A 1.398(12) C9A C10A 1.405(12) C9A C13A 1.485(13)C10A C11A 1.339(13) C12A C14A 1.497(12) C14A C15A 1.527(16) C14A C16A1.541(14) O3S C12S 1.51(3) O3S C15S 1.44(2) C11S C12S 1.64(5) C11T C12S1.51(3) C12S C13S 1.49(5) C12S C13T 1.39(5) C13S C14S 1.70(5) C13T C14S1.55(5) C14S C15S 1.39(3) O2S C7S 1.43(2) O2S C10S 1.52(3) C6S C7S1.38(3) C7S C8S 1.54(4) C8S C9S 1.57(4) C9S C10S 1.34(3) O1S C2S1.429(18) O1S CSS 1.41(2) C1S C2S 1.45(3) C2S C3S 1.46(3)

TABLE 6-4 Bond Angles in for COMPOSITION 4A. Atom Atom Atom Angler C5CO1C C1C 116.5(6) C12C O3C C2C 117.1(7) O1C C1C C2C 105.3(6) O1C C1C C3C108.6(7) C3C C1C C2C 111.5(7) O3C C2C C1C 106.6(6) O3C C2C C4C 108.9(6)C1C C2C C4C 110.9(7) O5C C3C C1C 109.9(7) O6C C3C O5C 126.1(8) O6C C3CC1C 124.0(8) O7C C4C O8C 127.3(8) O7C C4C C2C 122.3(8) O8C C4C C2C110.4(7) O1C C5C C6C 110.8(8) O2C C5C O1C 124.1(8) O2C C5C C6C 125.1(8)C7C C6C C5C 119.6(9) C11C C6C C5C 121.9(9) C11C C6C C7C 118.1(10) C8CC7C C6C 122.4(12) C9C C8C C7C 119.1(13) C10C C9C C8C 120.6(12) C9C C10CC11C 119.5(13) C6C C11C C10C 119.7(12) O3C C12C C13C 112.4(8) O4C C12CO3C 122.7(8) O4C C12C C13C 124.9(8) C14C C13C C12C 120.6(9) C18C C13CC12C 120.4(9) C18C C13C C14C 118.8(10) C15C C14C C13C 120.1(10) C16CC15C C14C 120.5(10) Cl5C C16C C17C 118.7(12) C16C C17C C18C 120.4(14)C13C C18C C17C 120.5(12) C1B N1B C8B 115.5(7) C2B N1B C1B 121.7(7) C2BN1B C8B 122.6(7) C3B N2B C2B 116.6(7) C1B N3B C7B 126.9(8) C12B N4B CHB118.8(8) O1B C1B N1B 121.2(8) O1B C1B N3B 122.4(9) N3B C1B N1B 116.3(8)N1B C2B C6B 119.7(8) N2B C2B N1B 117.3(7) N2B C2B C6B 123.1(8) N2B C3BC11B 116.9(7) N2B C3B C4B 125.0(8) C4B C3B C11B 118.0(7) F1B C4B C3B121.5(8) F1B C4B C5B 120.2(8) CSB C4B C3B 118.3(8) C4B CSB C6B 119.2(8)C2B C6B C7B 120.9(8) C5B C6B C2B 117.7(8) C5B C6B C7B 121.2(8) O2B C7BN3B 121.3(9) O2B C7B C6B 124.9(9) N3B C7B C6B 113.8(7) C9B C8B N1B118.0(7) C12B C8B N1B 120.2(7) C12B C8B C9B 121.7(7) C8B C9B C13B121.4(8) C10B C9B C8B 116.6(8) C10B C9B C13B 122.0(8) C11B C10B C9B119.9(8) N4B C11B C10B 123.0(8) N4B C12B C8B 119.9(8) N4B C12B C14B116.2(7) C8B C12B C14B 123.8(7) C12B C14B C 15B 113.0(8) C12B C14B C16B110.5(8) C15B C14B C16B 111.7(9) C1A N1A C2A 121.5(7) C1A N1A C8A118.9(7) C2A N1A C8A 119.5(7) C3A N2A C2A 116.6(8) C7A N3A C1A 126.9(8)C12A N4A C11A 120.6(7) O1A C1A N1A 120.7(8) O1A C1A N3A 122.4(8) N3A C1AN1A 116.9(8) N2A C2A N1A 116.6(7) N2A C2A C6A 124.3(8) C6A C2A N1A119.1(8) N2A C3A C11A 116.6(7) N2A C3A C4A 123.7(9) C4A C3A C11A119.7(8) F1A C4A C3A 119.7(9) C5A C4A F1A 121.1(9) C5A C4A C3A 119.2(9)C4A CSA C6A 119.7(9) C2A C6A C5A 116.4(9) C2A C6A C7A 121.0(8) C5A C6AC7A 122.6(8) O2A C7A N3A 120.9(9) O2A C7A C6A 124.4(9) N3A C7A C6A114.7(8) C9A C8A N1A 118.0(7) C9A C8A C12A 122.2(7) C12A C8A N1A119.7(7) C8A C9A C10A 115.9(8) C8A C9A C13A 123.3(8) C10A C9A C13A120.7(8) C11A C10A C9A 119.5(8) C10A C11A N4A 123.5(8) N4A C12A C8A118.2(8) N4A C12A C14A 118.8(7) C8A C12A C14A 123.0(7) C12A C14A C15A110.2(8) C12A C14A C16A 109.8(8) C15A C14A C16A 111.6(9) C15S O3S C12S101.0(16) O3S C12S C11S 98(2) C11T C12S O3S 114(3) C13S C12S O3S 101(3)C13S C12S C11T 119(4) C13T C12S O3S 108(3) C13T C12S C11S 124(4) C12SC13S C14S 94(3) C12S C13T C14S 106(3) C15S C14S C13S 105(2) C15S C14SC13T 98(2) C14S C15S O3S 109.6(18) C7S O2S C10S 109.8(17) O2S C7S C8S98(2) C6S C7S O2S 110(2) C6S C7S C8S 105(2) C7S C8S C9S 108(3) C10S C9SC8S 104(3) C9S C10S O2S 110(2) C5S O1S C2S 99.3(14) O1S C2S C1S115.3(16) O1S C2S C3S 109.3(14) C1S C2S C3S 113.3(18) C2S C3S C4S102.4(19) C3S C4S C5S 100(2) O1S C5S C4S 110(2)

TABLE 6-5 Hydrogen Fractional Atomic Coordinates (×10⁴) and EquivalentIsotropic Displacement Parameters (Å² × 10³) for COMPOSITION 4A. U_(eq)is defined as 1/3 of the trace of the orthogonalised U_(ij). Atom x y zU_(eq) H5C 2738 5676 7079 75 H8C 7257 4612 3523 74 H1C 5642 5286 5648 49H2C 4246 5000 4813 47 H7C 8030 3133 7918 117 H8CA 8829 1329 8674 113 H9C8046 −520 8781 129 H10C 6325 −589 8302 117 H11C 5582 1286 7440 113 H14C824 2153 5456 67 H15C −125 131 6310 90 H16C 854 −1479 7197 118 H17C 2922−1070 6936 117 H18C 3724 966 6375 104 H3B 6442 7830 7268 59 H5B 42843988 9467 59 H10B 10491 6619 10136 58 H11B 12147 6175 8883 53 H13A 76377233 9947 82 H13B 8351 6990 10789 82 H13C 7521 5873 10864 82 H14B 91415177 7246 64 H15A 11415 4105 7168 98 H15B 10492 3806 6645 98 H15D 100553410 7957 98 H16A 10303 7074 5885 109 H16B 10596 6013 5392 109 H16D11573 6448 5797 109 H3A 14191 5912 1268 55 HSA 14875 1914 3171 65 H10A9147 3010 831 57 H11A 7601 3622 2016 54 H13D 12178 3715 8 83 H13E 112362674 104 83 H13F 11970 2347 938 83 H14A 10773 4660 3501 60 H15E 98176386 3737 106 H15F 8738 6107 3360 106 H15G 10127 6543 2480 106 H16E 81743966 4816 96 H16F 9227 4126 5267 96 H16G 9198 3006 4874 96 H11D 403 12111721 212 H11E -600 −1054 12212 212 H11F -968 318 11703 212 H11G −1060−259 11882 290 H11H −1114 −1599 11825 290 H11I −1750 −509 11154 290 H12S128 −753 9931 215 H12A -557 −686 10345 215 H13G 1497 −1393 10872 182H13H 1451 −115 11089 182 H131 1087 −1267 9548 182 H13J 1458 −1412 10592182 H14D 3019 696 9168 186 H14E 2460 −284 8839 186 H14F 2628 537 9673186 H14G 2896 104 8645 186 H15H 1338 997 8243 167 H15I 1947 1996 8512167 H6SA 6575 −2283 5108 218 H6SB 5393 −1922 4756 218 H6SC 5233 −24176034 218 H7S 5282 −222 5136 186 H8SA 7003 307 3490 290 H8SB 7680 −8614007 290 H9SA 8880 527 4152 184 H9SB 7923 1561 4003 184 H10D 7155 10555667 243 H10E 8144 66 5812 243 H1SA 6626 7716 2249 264 H1SB 5541 84962685 264 H1SC 6226 8839 1383 264 H2S 5277 6457 2441 126 H3SA 3789 75063528 175 H3 SB 3293 6455 3231 175 H4SA 3168 9048 2302 249 H4SB 2364 79652275 249 H5SA 3745 8410 500 195 H5SB 4689 9246 647 195

TABLE 6-4 Hydrogen Bond information for COMPOSITION 4A. D H A d (D-H)/Åd (H-A)/Å d (D-A)/Å D-H-A/deg O5C H5C N4B¹ 0.84 1.82 2.656 (9) 170.9 O8CH8C N4A 0.84 1.79 2.624 (9) 171.6 N3B H3B O2S² 0.88 1.94 2.805 (12)168.1 C14B H14B N1B 1.00 2.44 2.937 (12) 110.1 N3A H3A O1S³ 0.88 1.952.798 (12) 160.7 ¹−1 + x, + y, + z; ²+ x, 1 + y, + z; ³1 + x, + y, + z

TABLE 6-5 Atomic Occupancies for all atoms that are not fully occupiedin COMPOSITION 4A. Atom Occupancy C11S 0.5 H11D 0.5 H11E 0.5 H11F 0.5CUT 0.5 H11G 0.5 H11H 0.5 H11I 0.5 H12S 0.5 H12A 0.5 C13S 0.5 H13G 0.5H13H 0.5 C13T 0.5 H13I 0.5 H13J 0.5 H14D 0.5 H14E 0.5 H14F 0.5 H14G 0.5

The foregoing is merely illustrative of the invention and is notintended to limit the invention to the disclosed uses. Variations andchanges, which are routine to one skilled in the art, are intended to bewithin the scope and nature of the invention, which are defined in theappended claims. All mentioned references, patents, applications andpublications, are hereby incorporated by reference in their entirety, asif here written.

What is claimed is:
 1. A composition, the composition comprising acompound of Formula 4:

and a compound of Formula B:


2. The composition of claim 1, wherein the compound of Formula 4 is acompound of Formula 5M:


3. The composition of claim 1, wherein the compound of Formula 4 is acompound of Formula 5P:


4. The composition of claim 1, wherein the compound of Formula B is acompound of formula B1:


5. The composition of claim 1, wherein the compound of Formula B is acompound of formula B2:


6. The composition of claim 1, wherein the composition comprises a 2 to1 ratio of the compound of Formula 4 to the compound of Formula B. 7.The composition of claim 1, wherein the composition further comprises2-methyltetrahydrofuran having the formula:


8. The composition of claim 1, wherein the ratio of the2-methyltetrahydrofuran to the compound of formula B is 2 to
 1. 9. Thecomposition of claim 1, wherein the composition has the formula:


10. The composition of claim 9, wherein the composition has the formula:


11. The composition of claim 9, wherein the composition has the formula:


12. The composition of claim 9, wherein the composition has the formula:


13. The composition of claim 9, wherein the composition has the formula:


14. The composition of claim 1, wherein the composition is in acrystalline state.
 15. A method of making a composition of formula 4a,the method comprising reacting a compound 4, having the followingchemical structure:

with a compound B1, having the formula:

in the presence of 2-methyltetrahydrofuran to form the composition offormula 4a, having the structure:


16. A method of obtaining a compound of formula 5M, having the followingchemical structure:

the method comprising: a) reacting a compound 4, having the followingchemical structure:

with a compound B1, having the formula:

in the presence of 2-methyltetrahydrofuran to form a composition offormula 4a, having the structure:

as crystals; b) isolating composition 4a, and c) treating the isolatedcomposition 4a with a base to produce the compound of formula 5M. 17.The method according to claim 16, wherein the base is Na₂HPO₄.
 18. Themethod according to claim 16, wherein the base is NaHCO_(3.)
 19. Acomposition, the composition comprising a compound of Formula 4:

and a compound of Formula 11:


20. The composition of claim 19, wherein the compound of Formula 4 is acompound of Formula 5M:


21. The composition of claim 19, wherein the compound of Formula 4 is acompound of Formula 5P:


22. The composition of claim 19, wherein the compound of Formula 11 is acompound of formula 11 a:


23. The composition of claim 19, wherein the compound of Formula 11 is acompound of formula 11b:


24. The composition of claim 19, wherein the composition has theformula:


25. The composition of claim 19, wherein the composition has theformula:


26. The composition of claim 19, wherein the composition has theformula:


27. The composition of claim 19, wherein the composition has theformula:


28. The composition of claim 19, wherein the composition comprises a 1to 1 ratio of the compound of Formula 4 to the compound of Formula 11.29. The method of claim 16, wherein the compound of formula 5M is usedas an intermediate to generate a compound having the Formula 9:


30. The method of claim 29, wherein the method further comprises mixingthe compound of Formula 9 with at least one pharmaceutically acceptableexcipient to form a pharmaceutical composition.